Biosensor performance indicator for intraoral appliances

ABSTRACT

Methods and apparatuses for monitoring either or both the performance of an orthodontic appliance for repositioning a patient&#39;s teeth and/or the user compliance in wearing the appliance using a biosensor. The apparatuses described herein may include one or more sensors, including biosensors, electrical sensors, or both, configured to generate sensor data related to user compliance and/or repositioning of the patient&#39;s teeth by an orthodontic appliance.

CROSS REFERENCE TO RELATED APPLICATIONS

This patent application claims priority to U.S. Provisional PatentApplication No. 62/525,082, filed Jun. 26, 2017, titled “BIOSENSORPERFORMANCE INDICATOR FOR INTRAORAL APPLIANCES,” which is hereinincorporated by reference in its entirety.

INCORPORATION BY REFERENCE

All publications and patent applications mentioned in this specificationare herein incorporated by reference in their entirety to the sameextent as if each individual publication or patent application wasspecifically and individually indicated to be incorporated by reference.

FIELD

Orthodontic appliances, including dental appliances, including shellaligners, with one or more sensors for monitoring use and/or progress ofthe appliance.

BACKGROUND

Orthodontic procedures typically involve repositioning a patient's teethto a desired arrangement in order to correct malocclusions and/orimprove aesthetics. To achieve these objectives, orthodontic appliancessuch as braces, shell aligners, and the like can be applied to thepatient's teeth by an orthodontic practitioner. The appliance can beconfigured to exert force on one or more teeth in order to effectdesired tooth movements according to a treatment plan.

During orthodontic treatment with patient-removable appliances, thepractitioner may rely on the patient to comply with the prescribedappliance usage. In some instances, a patient may not wear theorthodontic appliance as prescribed by the practitioner. Extendedremoval of the appliance, for any reason beyond what is recommended, mayinterrupt the treatment plan and lengthen the overall period oftreatment.

In some instances, the forces that are actually applied to a patient'steeth by an orthodontic appliance may differ from the intended forcesfor treating the teeth. Discrepancies between the planned and achievedrepositioning forces may result in incomplete or undesirable toothmovements and deviations from the prescribed treatment plan.Accordingly, improved approaches for monitoring orthodontic applianceperformance, treatment progress, and patient compliance are needed.

SUMMARY OF THE DISCLOSURE

In general, described herein are methods and apparatuses for detectingone or more biomarkers from within a subject's oral cavity. Theapparatuses (including devices and systems) described herein may includeany oral appliance, including (but not limited to) aligners, palatalexpanders, retainers, and the like.

For example, described herein are methods and apparatuses (includingsystems and devices) for monitoring one or more of: the performance ofan orthodontic appliance for repositioning a patient's teeth, thepatient's physiological condition and/or the user compliance in wearingthe appliance. The apparatuses described herein may include one or moresensors, including biosensors, electrical sensors, or both, configuredto generate sensor data related to user compliance and/or repositioningof the patient's teeth by an orthodontic appliance. For example, thedata can be indicative of patient wearing compliance, the amount oftooth movement achieved, the amount of force and/or pressure actuallyapplied to the teeth by the appliance, bone remodeling processes andstages, tissue health, bacterial activity in the oral cavity, or anycombination thereof. Advantageously, the embodiments described hereinprovide high value data that allows the practitioner to quantitativelyassess whether the orthodontic appliance is repositioning the patient'steeth as planned. The data can be used as feedback to adjust thepatient's treatment plan, also known as “adaptive closed-looptreatment,” and can also inform the design and planning of futureappliance based orthodontic procedures. Based on such data, a treatmentplan can be adaptively modified. For instance, to estimate whether andwhen the patient is ready for next stage Aligner.

This methods and apparatuses described herein may be used for monitoringand analysis of bioagents in the intraoral cavity while an intra-oralappliance (e.g., aligner, palatal expander, etc.) is in use. Theseapparatuses and methods may collect information (data), including dataabout tooth movement phases, via analysis of biomarkers in saliva orgingival crevicular fluid (GCF).

Also described herein are apparatuses for monitoring patient complianceand/or performance of an orthodontic appliance for repositioning apatient's teeth. These apparatuses may include an orthodontic appliance,such as an aligner comprising one or more teeth-receiving cavitiesshaped to reposition the patient's teeth from an initial arrangementtowards a target arrangement and one or more sensors, includingbiosensors, configured to generate sensor data. The sensor data may berelated to one or more biomarkers from saliva and/or GCF. The apparatusmay also comprise a processor configured to process the sensor data, toevaluate the performance of the orthodontic appliance in effecting therepositioning of the patient's teeth, patient health including oralhealth, and/or patient compliance.

Thus, an orthodontic appliance as described herein may include one ormore biosensors configured to obtain data indicative of a biomarker inthe patient's saliva, GCF, tissue or tissue (including tooth) surface,and one or more processors operably coupled to the biosensor(s) andconfigured to process the biosensor data so as to generate patientcompliance data (e.g., enabling electronic monitoring of patientcompliance with a prescribed course of orthodontic treatment), patienttreatment monitoring data, and/or patient health (e.g., oral health)data. Advantageously, these apparatuses and methods may increase improvetreatment efficacy and may provide patient data useful to thepractitioner for designing and monitoring orthodontic treatments.

Thus, described herein are devices for repositioning a patient's teethand monitoring a biomarker indicative of performance of the deviceand/or patient compliance in wearing the device. Any of these devicesmay include: an orthodontic appliance comprising one or more (e.g., aplurality of) teeth-receiving cavities shaped to reposition thepatient's teeth from an initial arrangement towards a targetarrangement; a biosensor comprising a bioreceptor material configured tospecifically interact with the biomarker and a biotransducer materialconfigured to transduced interaction of the bioreceptor with thebiomarker into an electrical signal; and a processor in electricalcommunication with the biosensor and configured to process theelectrical signal into sensor data to evaluate one or more of: theperformance of the orthodontic appliance in effecting the repositioningof the patient's teeth and patient compliance.

In general, the bioreceptor may comprise a protein that selectivelybinds to the biomarker. For example, the bioreceptor may include aprimary and/or a secondary protein, such or a portion of protein, suchas an antibody, antibody fragment, or the like, including antibody-basedmolecules, and/or enzymes. In general, any molecule that recognizes atarget molecule with specificity may be used. For example, thebioreceptor may be an enzyme that selectively acts on the biomarker.

Any biomarker may be targeted, particularly but not exclusively abiomarker present in saliva. Alternatively, the biomarker may be abiomarker in the blood, gingiva, or teeth. In some cases, the biomarkermay be a biomechanical property, such as heart rate, blood pressure, orthe like. Biomarkers that may be present in saliva may include one ormore of: Calgranulin-B, Serum albumin precursor, Immunoglobulin J chain,Ig alpha-1 chain C region, Cysteine-rich secretory protein 3 precursor(CRISP-3), Hemoglobin subunit beta, Stratifin, and soluble RANK Ligand(sRANKL). The biomarker may be present in gingival crevicular fluid(GCF). For example, the biomarker may be one or more of: prostaglandinE2, Substance P, epidermal growth factor, transforming growth factor,Receptor activator of nuclear factor kappa-B ligand (RANKL), Granulocyetmacrophage colony stimulation factor, α2 microglobulin, Interleukin 1β,Myeloperoxidase, hyaluronic acid and Chondroitin sulfate.

Further, any appropriate biosensor may be used, including electrodes.For example, the biosensor may comprise an electrode made nanowiresand/or nanoparticles. Carbon nanotubes, graphene, noble metal-basednanoparticles and the like can be used to form all or portion of thebiosensor(s) herein, for example, by forming a conductive trace. Thesematerials may be patterned, including forming onto an orthodontic deviceas a flexible trace, e.g., using a printed or otherwise appliedconductive ink. The conductive ink may include a material (such as anelastomeric binder) to increase flexibility. For example, the biosensormay comprise a conductive ink and an elastomeric binder such as one ormore of: silicone, fluorine rubber, polyurethane and isoprene blockco-polymers. Flexible electrodes/conductive traces may be used as partof the biosensor and as connections between the biosensor and any othercomponents, including the processor. Examples of conductive ink include:activate carbon knitted carbon fibers (with a binder such as, e.g.,polytetrafluoroethylene as a), RhO2/SWCNT (with a binder such as, e.g.,sodium dodecyl sulfate), graphene (with a binder such as, e.g., ethylcellulose), Ketjenblack porous carbon (with a binder such as, e.g., astyrene-butadiene rubber), PEG coated silver flakes (with a binder suchas, e.g., a polyurethane), Ag/AgCl ink and/or multi-walled carbonnanotube (MWCNT) and silver (Ag) nanoparticles (with a binder such as,e.g., a polyurethane), silver flakes or silver particles (with a bindersuch as, e.g., a binder such as, e.g., copolymer fluoroelastomer ofvinylidene-fluoride/Hexfluoropropylene Zonyl-FS 300 fluorosurfactantand/or hexyl acetate capsules with organic solvent (hexyl acetate)acrylic).

The biosensor may include a conductive substrate to which thebiotransducer is attached and an insulating layer.

In general, any of these devices may include a reference electrodecomprising the biotransducer material but not a bioreceptor material.Further, any of these devices may include a plurality of biosensorsoperably coupled to different portions of the orthodontic appliance.

Any of these devices may include a second (or more) sensor operativelycoupled to the orthodontic appliance, wherein the additional sensor(s)comprises one or more of: a force or pressure sensor, a temperaturesensor, or a movement sensor.

In general, the biosensor may be integrated with the orthodonticappliance, coupled to a tooth, or a combination thereof. Any of thesedevices may include a communication module configured to transmit thesensor data to a remote device.

Any of the apparatuses described herein may include one or moremicroneedles, including arrays of microneedles. See, e.g.,PCT/US2012/053544 (herein incorporated by reference in its entirety) forexamples of microneedles that may be used in any of the apparatusesdescribed herein. In general, these apparatuses (and methods of usingthem) may include a mechanism such a microneedle for drawing anddetecting any of the analytes described herein. For example, an array ofhollow probes (e.g., tubes, needles, etc.) can extract biomaterialtransdermally. In some variations the probes may include amperometric,voltamemertric, or potentiometric sensors within the probe to measureanalytes in the sample.

Also described herein are methods of monitoring an orthodontic device ina subject's oral cavity. These methods may be used to modify a dentaltreatment plan (e.g., an orthodontic treatment plan). Also, in general,any of these methods may be used for monitoring oral health, generally,regardless of the performance of the aligner. Thus, any of these methodsmay be configured as methods of monitoring a subject's health (or oralhealth) using any of the devices and methods described herein.

For example, a method of monitoring an orthodontic device in a subject'soral cavity may include: inserting an orthodontic appliance comprisingone or more teeth-receiving cavities shaped to reposition the patient'steeth from an initial arrangement towards a target arrangement onto thepatient's teeth; detecting a biomarker within the subject's saliva orgingival crevicular fluid using a biosensor configured to specificallyinteract with the biomarker coupled to the orthodontic appliance; andprocessing signals from the biosensor to evaluate one or more of: theperformance of the orthodontic appliance in effecting the repositioningof the patient's teeth and patient compliance.

Detecting the biomarker may include detecting the biomarker within thesubject's saliva. For example, detecting the biomarker may includedetecting at least one of: Calgranulin-B, Serum albumin precursor,Immunoglobulin J chain, Ig alpha-1 chain C region, Cysteine-richsecretory protein 3 precursor (CRISP-3), Hemoglobin subunit beta,Stratifin, and soluble RANK Ligand (sRANKL). Detecting the biomarker maycomprises detecting the biomarker within the subject's gingivalcrevicular fluid; for example, detecting the biomarker may comprisedetecting at least one of: prostaglandin E2, Substance P, epidermalgrowth factor, transforming growth factor, Receptor activator of nuclearfactor kappa-B ligand (RANKL), Granulocyet macrophage colony stimulationfactor, α2 microglobulin, Interleukin 1β, Myeloperoxidase, hyaluronicacid and Chondroitin sulfate. Detecting the biomarker may comprisedetecting an interaction between the biomarker and a bioreceptor of thebiomarker that is transduced by a biotransducer of the biosensor into anelectrical signal.

In general, processing signals from the biosensor may include wirelesslytransmitting the signals, and/or transmitting them via a wiredconnection.

As mentioned, any of these methods may be configured to modify a patienttherapy based on the output of the sensing, including modifying atreatment plan (e.g., orthodontic treatment plan). For example,processing the signals may comprise modifying an orthodontic treatmentplan for the patient based on the signals. Alternatively, any of thesemethods may include modifying the treatment plan. Modifying theorthodontic treatment plan may comprise generating a second orthodonticappliance comprising one or more teeth-receiving cavities shaped toreposition the patient's teeth from a current arrangement towards asecond target arrangement. Modifying the orthodontic treatment plan maycomprise removing the orthodontic appliance.

Also described herein are systems in which the aligner include an opencircuit that is closed when the patient wears the aligner on theirteeth. For example, described herein are systems for repositioning apatient's teeth and monitoring a biomarker indicative of performance ofan orthodontic appliance and/or patient compliance in wearing theappliance, the system comprising: the orthodontic appliance comprisingone or more (e.g., a plurality of) teeth-receiving cavities shaped toreposition the patient's teeth from an initial arrangement towards atarget arrangement; a first electrical contact within theteeth-receiving cavity of the orthodontic appliance; a second electricalcontact within the teeth-receiving cavity of the orthodontic appliance;an electrical circuit coupled to the first electrical contact and thesecond electrical contact, wherein the electrical circuit is open untilthe orthodontic appliance is worn on the patient's teeth so that anelectrical trace on the patient's tooth or teeth couples to both firstelectrical contact and the second electrical contact to close theelectrical circuit; and a processor in electrical communication with theelectrical circuit and configured to generate a signal when theelectrical circuit between the first electrical contact and the secondelectrical contact is closed.

Any of these systems may include an electrically conductive traceconfigured to be mounted on the patient's teeth, the electricallyconductive trace having a first node and a second node, wherein thefirst node is configured to contact the first electrical contact and thesecond node is configured to contact the second electrical contact.

These systems may also include one or more sensors coupled to theelectrical circuit, wherein the sensor is configured to operate when theelectrical circuit is closed.

These systems may include a power source on the orthodontic appliance.The power source may be a battery or the like, particularly arechargeable battery. In any of the devices and systems describedherein, the power source may include a biofuel cell sensor that ispowered using metabolites in saliva as fuel as well as as analytes.

Any of these systems and devices may include a processor, and theprocessor may be configured to wirelessly transmit the signal. Theprocessor may comprise a data logger to record the signals. Theprocessor may be configured to periodically transmit the signal whilethe electrical circuit is closed.

Also described herein are methods of determining patient compliance inwearing an orthodontic appliance that comprises one or moreteeth-receiving cavities shaped to reposition the patient's teeth froman initial arrangement towards a target arrangement. For example, themethod may include: closing an electrical circuit on the orthodonticappliance by placing the orthodontic appliance on the patient's teethand electrically coupling a pair of electrical contacts within theteeth-receiving cavities with a first node and a second node of anelectrical trace on the patient's teeth, so that the electrical tracecloses the electrical circuit; and emitting a signal when the electricaltrace closes the electrical circuit indicating compliance.

Any of these methods may include storing the signal, and/or periodicallyemitting signals while the electrical trace closes the electricalcircuit. Emitting the signal may comprise wirelessly transmitting thesignal. The methods may also include logging the signal to track patientcompliance.

For example, described herein are removable orthodontic devicesincluding a biosensor. In some variations, the biosensor may be abiosensor system that is configured to detect a biomarker that indicatesa phase of movement of the patient's teeth during an orthodontisttreatment. For example, the biosensor may be configured to detect one ormore biomarkers, such as one or more salivary proteins, the expressionof which changes during orthodontic treatment. As a non-limitingexample, protein S100-A9 (S100 calcium-binding protein A9 orCalgranulin-B), serum albumin precursor, immunoglobulin J chain,immunoglobulin J chain, and Ig alpha-1 chain C region each showdown-regulation two weeks (fourteen days) following the start of anorthodontic treatment. These markers may be in saliva or in GCF.Similarly, expression of IL-6, IL-8 levels in gingival crevicular fluidmay be altered after force application is applied and remodeling hasbegun. The levels of markers for inflammation, remodeling and/or enzymesassociated with bone resorption, formation, cell necrosis, collagenremodeling, etc. may be detected and may be associated with the phase oftooth movement.

Tooth movement induced by orthodontic force application may becharacterized by remodeling changes in the dental and periodontaltissues and may include deflection, or bending, of the alveolar bone andremodeling of the periodontal tissues, including the dental pulp,periodontal ligament (PDL), alveolar bone, and gingiva. The appliedforce causes the compression of the alveolar bone and the PDL on oneside, while on the opposite side the PDL is stretched.

Orthodontic tooth movement may be described as having three or morephases. For example, tooth movement may have an initial phase, a lagphase, and a post-lag phase. The initial phase may be characterized byimmediate and rapid movement and may occurs 24 hours to 48 hours afterthe first application of force (above a movement threshold) to thetooth. This rate is largely attributed to the displacement of the toothin the PDL space. The lag phase may typically last for 20 to 30 days,depending on the amount of force applied, and may show relatively littleto no tooth displacement. This phase may be marked by PDL hyalinisationin the region of compression. Subsequent tooth movement typically doesnot occur until the cells complete the removal of all of the necrotictissues. The postlag phase follows the lag phase, during which the rateof movement may again increase.

Thus, a removable orthodontic devices including a biosensor may include:a plurality of tooth receiving cavities configured to receive aplurality of teeth and to exert one or more orthodontic repositioningforces on the plurality of teeth; and at least one biosensor systemcomprising: a bioreceptor configured to cause a first interaction withone or more tooth motion biomarkers in fluid in the oral cavity, thefirst interaction being related to a first biomarker expression changeof the one or more tooth motion biomarkers, and the first biomarkerexpression change being associated with a specific phase of toothmovement of the plurality of teeth; and a biotransducer coupled to thetooth motion bioreceptor, the biotransducer configured to transduce thefirst interaction into a first interaction signal representative of thefirst interaction.

The specific phase of tooth movement may correspond to a velocity ofroot movement of roots of the plurality of teeth. For example, the levelof the biomarker may reflect the velocity of the root movement. In somevariations, if the velocity of root movement is substantially zero, thebiosensor may indicate that a particular portion of an orthodontictreatment plan may be terminated, and a new stage (e.g., a new aligner,such as the next aligner in a sequence) may be applied.

Additional biomarkers may be used, including biomarkers that reflectchanges indicative of other stages of tooth movement and/or of thepresence and/or absence of the apparatus in the patient's mouth (e.g.,compliance). For example, the bioreceptor may be configured to cause asecond interaction with one or more compliance biomarkers, the secondinteraction being related to a second biomarker expression changeassociated with compliance by the patient; and the biotransducer may beconfigured to transduce the second interaction into a second interactionsignal representative of the second interaction.

Any of the biosensor systems described herein may include one or moreprocessor configured to process the first interaction signal into sensordata used to provide one or more recommendations to change the removableorthodontic appliance at a specified time. The one or more processorsmay monitor the level of the biomarkers over time, and may store,analyze and/or transmit these levels. In some variation the one or moreprocessors may compare the signal strength to a threshold; when thebiomarker signal falls above (or in some variations, below, depending onthe biomarker), the processor may indicate that the dental applianceshould be removed and/or changed. As mentioned above, the at least onebiosensor system may include one or more of: a memory, a power source, acommunication unit, and an antenna.

Any of these systems may include a biosensor housing formed from atleast a portion of the plurality of tooth receiving cavities and beingconfigured to physically couple the bioreceptor to at least one tooth ofthe plurality of tooth. The bioreceptor may include a protein thatselectively binds to the tooth motion biomarker (e.g., an antibody orantibody fragment, enzyme, etc.). For example, the bioreceptor mayinclude a protein (e.g., one or more of a primary and/or a secondaryprotein). The bioreceptor may comprises an enzyme that selectively actson the tooth motion biomarker.

The fluid may be saliva, gingival cervicular fluid (GCF), etc. In somevariations, the tooth movement biomarker is one or more of:Calgranulin-B, Serum albumin precursor, Immunoglobulin J chain, Igalpha-1 chain C region, Cysteine-rich secretory protein 3 precursor(CRISP-3), Hemoglobin subunit beta, Stratifin, and soluble RANK Ligand(sRANKL). The removable orthodontic device of claim 1, wherein the fluidcomprises gingival crevicular fluid (GCF). In some variations, the toothmovement biomarker is one or more of: prostaglandin E2, Substance P,epidermal growth factor, transforming growth factor, Receptor activatorof nuclear factor kappa-B ligand (RANKL), Granulocyet macrophage colonystimulation factor, α2 microglobulin, Interleukin 1β, Myeloperoxidase,hyaluronic acid and Chondroitin sulfate.

In any of the apparatuses (e.g., removable orthodontic devices, etc.)described herein, the apparatus may gather a baseline level of thebiomarker. For example, the a baseline may be detected from the patientby having the patient wear a first removable orthodontic device that isnot configured to move the teeth prior to wearing an orthodontic devicethat is configured to move the teeth. The first (baseline gatheringdevice) may be worn for any appropriate time (e.g., 5 minutes or more, 1hour or more, 2 hours or more, 3 hours or more 4 hours or more, sixhours or more, 12 hours or more, 1 day or more, 3 days or more, one weekor more, two weeks or more, etc.) for some minimum amount of time (5minutes, 1 hour, 2 hours, 3 hours, 4 hours, 6 hours, 12 hours, 1 day, 3days, 1 week, 2 weeks, etc.). This first device may be configured torest on the teeth without applying any substantial moment (force) tomove the teeth. The biosensor(s) on the device may therefore collectpatient-specific baseline information that may be stored, analyzedand/or transmitted for use in the subsequent appliances (e.g., removableorthodontic devices) that are configured to move the teeth. For examplein some variations, a series of appliances (removable orthodonticdevices, e.g., aligners) may be worn in a sequence to move the teeth.Each subsequently-worn appliance that includes a biosensor system maycollect the baseline information (e.g., baseline level of the one ormore biomarker) by receiving this information collected from the initialdevice. This information may be transmitted to the subsequent removableorthodontic device(s) by the initial device and/or by a remote processorthat communicates with the removable orthodontic device (e.g.,wirelessly). Alternatively or additionally, the baseline information maybe transmitted and stored remotely and comparisons to the baseline maybe made remotely as well.

In some variations the apparatus (e.g., the removable orthodonticdevice) may include a biosensor reference. For example, the at least onebiosensor system may include a reference electrode comprising thebiotransducer material but not a bioreceptor material.

In some variations the biosensor comprises one or more of nanowires andnanoparticles. In some variations, the biosensor comprises a conductiveink and an elastomeric binder. The elastomeric binder may include:silicone, fluorine rubber, polyurethane and isoprene block co-polymers.The biosensor may include a conductive substrate to which thebiotransducer is attached and an insulating layer.

Some of the apparatuses (including a removable orthodontic device) mayhave multiple biosensors thereon. For example, in some variations the atleast one biosensor system is one of a plurality of biosensor systemsoperably coupled to different portions of the removable orthodonticdevice.

Any of these apparatuses (e.g., removable orthodontic devices) describedherein may also include a physical sensor operatively coupled to theremovable orthodontic device, wherein the physical sensor is configuredto sense one or more of a force, a pressure, a temperature, or amovement of the plurality of teeth.

As mentioned above, any of these methods may also include acommunication module configured to transmit (e.g., wirelessly) thesensor data to a remote device.

The methods described herein may also include a method comprising:receiving a plurality of teeth in an oral cavity of a patient with aplurality of tooth receiving cavities of a removable orthodonticaligner, the plurality of tooth receiving cavities configured to exertone or more orthodontic repositioning forces on the plurality of teeth;placing a bioreceptor in contact with fluid in the oral cavity to causea first interaction with one or more tooth motion biomarkers in thefluid, the first interaction being related to a first biomarkerexpression change of the one or more tooth motion biomarkers, and thefirst biomarker expression change being associated with a specific phaseof tooth movement of the one or more teeth; transducing the firstinteraction into a first interaction signal representative of the firstinteraction; and providing the first interaction signal.

A removable orthodontic device as described herein may include: meansfor receiving a plurality of teeth in an oral cavity of a patient and toexert one or more orthodontic repositioning forces on the plurality ofteeth; and at least one biosensor system comprising: means for causing afirst interaction with one or more tooth motion biomarkers in fluid inthe oral cavity, the first interaction being related to a firstbiomarker expression change of the one or more tooth motion biomarkers,and the first biomarker expression change being associated with aspecific phase of tooth movement of the plurality of teeth; and meansfor transducing the first interaction into a first interaction signalrepresentative of the first interaction.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1A illustrates a removable orthodontic device (e.g., a toothrepositioning appliance, or aligner), configured as a shell havingcavities for fitting over teeth. FIGS. 1B-1D illustrate a series ofremovable orthodontic devices (configured as a tooth repositioningsystem) that may be worn over a subject's teeth to reposition them. Anyof these removable orthodontic devices may be configured as describedherein to include or operate in conjunction with the biosensors andsystems including biosensors as described herein.

FIG. 2A illustrates a method of orthodontic treatment using a pluralityof removable orthodontic devices (e.g., a series of appliances) such asthose shown in FIGS. 1B-1D. As mentioned, this method may be modified byusing feedback from the one or more biosensors (biosensor systems) asdescribed herein.

FIG. 2B schematically illustrates a method for monitoring performance ofan orthodontic appliance for repositioning a patient's teeth.

FIG. 3A schematically illustrates one example of an apparatus includinga monitoring device (e.g., electronics) that may include a biosensor ormay communicate with a biosensor coupled to the oral cavity (e.g.,teeth, gingiva, etc.). In FIG. 3A a biosensor is shown coupled orintegrated with the apparatus.

FIG. 3B schematically illustrates another example of an apparatusincluding a biosensor.

FIG. 4A illustrates a schematic of a biosensor apparatus (e.g., whichmay be part of a monitoring device) with an activation mechanism.

FIG. 4B illustrate schematically an example of a monitoring device 400with an activation mechanism.

FIG. 4C schematically illustrates another example of an apparatus (e.g.,including a monitoring device) configured as a biosensor with anactivation mechanism.

FIG. 4D is an example of an apparatus with an activation mechanismsimilar to that shown in FIGS. 4A and 4C. In this example, the apparatusincludes bio-receptors formed on a silicon nanowire atop a substrate.Nanoparticles and/or nanowires may be used as part of the bio-receptor.

FIG. 5A illustrates an example of a dental apparatus (e.g., a removableorthodontic device, also referred to as an orthodontic appliance)including an integrated monitoring device. FIG. 5B is a cross-sectionalview of the appliance of FIG. 5A.

FIG. 6A is an example of an apparatus including an electrical trace thatis bonded directly to the subject's teeth and configured to interactwith electrical circuitry and/or power on a wearable orthodontic piece(e.g., aligner). In this example, wearing the aligner properly on theteeth completes a circuit in the aligner that may accurately tracecompliance and/or may activate a sensor (e.g., biosensor). FIG. 6Billustrates the open circuit between the appliance (e.g., aligner, onleft) and conductive traces on teeth when the appliance is not worn onthe teeth or is improperly worn. FIG. 6C shows the closed circuit, whenthe appliance is worn so that the nodes on the teeth are coupled to thenodes on the appliance.

FIG. 7A is an example of a dental apparatus configured as a dentalaligner that includes a biosensor monitoring apparatus. In this example,the biosensor includes a swellable material that swells when binding toone or more biomolecules. Swelling (and therefore binding to thebiomarker(s) may be detected by detecting a pressure change in thematerial. FIGS. 7B and 7C schematically illustrate the biosensor portionof FIG. 7A, showing swelling of the swellable material (e.g., gel)before (FIG. 7B) and after (FIG. 7C) binding to the target biomarker.

FIG. 8 illustrates an example of a method of operating a removableorthodontic device (e.g., appliance) including a biosensor.

FIG. 9 illustrates an example of a method of modifying an orthodontictreatment plan using a removable orthodontic device including abiosensor.

FIG. 10A is an example of an orthodontic apparatus (e.g., aligner)including a biosensor including microfluidics for assaying one or morebiomarkers at different times.

FIG. 10B is a schematic illustration of one example of a biosensorincluding microfluidics and assay chambers that may be operated in achronological order to provide controlled sampling.

FIGS. 10C-10H illustrate the operation of a biosensor similar to thatshown in FIG. 10B, sampling over different time points.

FIG. 11 is example of another variation of an example of a biosensorincluding microfluidics and assay chambers that may be operated in achronological order to provide controlled sampling.

DETAILED DESCRIPTION

Described herein are methods and apparatuses for detecting a biomarkerin the intraoral cavity. In particular, these methods and apparatusesmay be configured to detect one or more biomarker, or a biomarker and aphysiological marker (such as body temperature, blood oxygenation,galvanic skin response) including, but not limited to, detection fromsaliva or gingival crevicular fluid (GCF).

For example, a generic apparatus as described herein may include asensor (e.g., biosensor) configured to be positioned in a subject's oralcavity and an electronics system in communication with the sensor, wherethe electronic system includes one or more of: a signal amplifier, asignal conditioner (e.g., filter, averaging, etc.), processor, memory(e.g., data logging unit), power source (e.g., battery, capacitor, etc.)and data communications (e.g., wired or wireless communications, such asBluetooth, Wifi, Zigbee, RFID, ultrasound, or the like). The sensortypically converts a biomarker detection into an electrical signal, andmay include a bioreceptor and a biotransducer. The bioreceptor isconfigured to interact with a specific analyte or subset of analytes andproduce a measurable signal (optical, chemical, mechanical, thermal,electrical, or some combination of these) that is transduced by thebiotransducer into an electrical signal that can be processed(amplified, filtered, averaged, combined, etc.) by the electronicsubsystem. The electronic subsystem may be an integrated system (e.g., aCMOS-based microprocessor). All or some of these components may be on orpart of an oral appliance, such as an aligner, brace, palatal expander,etc.

For example, FIG. 3A shows a portion of an apparatus that includes abiosensor (sensor(s) 306) comprising a bioreceptor and biotransducer, incommunication with an electronics system 335. The biosensor may beintegrated and/or incorporated with the electronics system 335 andconnected directly to a patient, etc., on a tooth, teeth, gingiva, etc.,or it may be connected to an oral appliance. The biosensor(s) 333 may beseparate from the electronics system 335 (indicated by the dashed lines.For example, the biosensor(s) may be part of a second oral applianceand/or directly mounted in the oral cavity (e.g., on a tooth/teeth), andmay couple to an oral appliance including all or some of the electronicssystem (e.g., battery/power source 316, processor 302, antenna 312,memory 304, etc.).

FIG. 3B is another example of a schematic illustrating a biosensorapparatus. In FIG. 3B, the biosensor(s) 356 are integrated with theelectronics system. The electronics system in this example include aprocessor (CPU 352) that may amplify, filter, analyze and/or store thesignals from the biosensor(s). The biosensor(s) may include abioreceptor and a biotransducer that converts interaction (e.g.,binding, enzymatic interaction, etc.) with a biomarker into anelectrical signal. The electronics may further include anoscillator/clock 358, and a memory (e.g., EEPROM 354). Additional memory(e.g., RAM 364) may also be included. The memory may store datagenerated by the biosensor(s) and/or control logic/command logic foroperating the apparatus. This logic may be modified (e.g.,programmable). Additional memory the electronics system (which may alsobe referred to herein as a sensor electronics system or sub-system) mayinclude communications circuitry (shown as an RFID front end 355) inFIG. 3B, which may also include or communicate with an antenna 357. Theelectronics system may also include power management circuitry 366, forregulating the power used by the electronics and/or biosensor(s). Thepower management circuitry may regulate a power source, such as abattery 367.

As schematically illustrated in FIG. 4A, a biosensors may include abioreceptor 411 that interacts with a biomarker and is functionallycoupled with a biotransducer 413. In this example, the bioreceptor isshown schematically including specific interaction sites that may engagewith a biomarker, e.g., by binding, enzymatically reacting with, etc.The biotransducer 413 is functionally linked to bioreceptor and maytransduce interaction between the biomarker and the bioreceptor into anelectrical signal (output) that is passed on to the electronic system(not shown, see, e.g., FIGS. 3A and 3B). For example, the biotransducermay bind to the bioreceptor and binding may be detected by thebiotransducer via an optical signal. In some variations, the bioreceptormay be bound to an optically transparent substrate through which lightmay be passed by the biotransducer; a change in the optical propertiesof the bioreceptor may correlate with binding of the biomarker to thebiotransducer. Alternatively, the bioreceptor may include a matrix(e.g., hydrogel) that interacts with the biomarker to modify a propertyof the matrix (e.g., electrical resistance, optical absorption,electrochemical potential, etc., and this modified property may bepolled and/or detected by the biotransducer. Other specific examples areprovided below.

FIGS. 4C and 4D also illustrate examples of biosensors that include abioreceptor 411 that interacts with a biomarker and is functionallycoupled with a biotransducer 413. In FIG. 4C, analyte 421 is detected byspecifically binding to a bioreceptor 411 (e.g., a molecularlyrecognizing material such as an enzyme, antibody, microorganism, Cdl,etc.) and this interaction is detected by the signal transductor formingthe biotransducer 413 (e.g., an electroactive substance detected by anelectrode, pH changing material detected by a pH electrode, anexothermic reaction being detected by a thermocouple, a photon-emittingreaction detected by a photon counter, a mass change detected by apiezoelectric device, etc.). Signals from the transducer are typicallyconverted to electrical signals (analog and/or digital) that are thentransmitted to a detector 423.

Any appropriate biomarker or biomarkers may be used. Biomarkers may bebiomolecules or byproducts of biomolecules that are present in the oralcavity (including saliva, GCF, and/or breath) and/or contaminants thatmay be present in the oral cavity, such as bacteria, yeast, etc.Biomolecules of particular interest include those that change inresponse to movement of the teeth due to an orthodontic procedure. Forexample, Table 1, below lists examples of biomarkers that may be testedusing the biosensor apparatuses described herein. For example in Table1, protein biomarkers include Protein S100-A9 (e.g., S100calcium-binding protein A9, Calgranulin-B), Serum albumin precursor,Immunoglobulin J chain, Ig alpha-1 chain C region, Cysteine-richsecretory protein 3 precursor (CRISP-3), Hemoglobin subunit beta(Hemoglobin beta chain, Beta-globin), and 14-3-3 protein σ (Stratifin,Epithelial cell marker protein 1).

TABLE 1 Summary of saliva proteins with expression change duringorthodontic treatment. (These are proteins that show statisticallysignificant change (P < 0.5) between day 0 and day 14 of orthodontictreatment.) Peptides matched/% Accession coverage Spot number MW(kDa)/pI (Protein Expression no. Protein name Known Function (UniProt)Theoretical Experimental score) change 1 Protein S100-A9 (i)Calcium-binding protein. P06702 13.2/5.71 14.5/5.47 4/24% (121) Down(S100 calcium- (ii) Promotes phagocyte regulated at binding proteinmigration and infiltration of day 14 A9) (Calgranulin- granulocytes atsites of B) wounding. (iii) Plays a role as a proinflammatory mediatorin acute and chronic inflammation. 2 Serum albumin (i) Good bindingcapacity P02768 69.3/5.92 14.8/7.00 8/11% (105) Down precursor forwater, Ca2+, Na+, K+, regulated at fatty acids, hormones, day 14bilirubin, and drugs. (ii) Main function is the regulation of thecolloidal osmotic pressure of blood. (iii) Major zinc transporter inplasma. 3 Immunoglobulin J (i) Serves to link two P01591 15.5/4.6234.0/3.86 10/30% (220)  Down chain monomer units of either regulated atIgM or IgA. day 14 (ii) Help to bind IgM or IgA to secretory component.4 Immunoglobulin J P01591 15.5/4.62 36.4/3.40 6/27% (198) Down chainregulated at day 14 5 Ig alpha-1 chain C (i) Major immunoglobulin P0187637.6/6.08 44.76/6.91  9/20% (260) Down region class in body secretions.regulated at (ii) Serve both to defend day 14 against local infectionand to prevent access of foreign antigens. 6 Cysteine-rich (i) Innateimmune response P54108 27.6/8.09 43.25/8.90  2/6% (94) Present secretoryprotein 3 (ii) Potential biological only at day precursor (CRISP- markerfor prostate cancer 14 3) 7 Hemoglobin Involved in oxygen transportP68871 15.9/6.75 14.83/9.50  6/27% (158) Present subunit beta from thelung to the various only at day 0 (Hemoglobin beta peripheral tissues.chain) (Beta- globin) 8 14-3-3 (i) Adapter protein. P31947 27.7/4.6844.76/4.20  5/22% (147) Present protein σ (Stratifin) (ii) Binds to alarge number only at day 0 (Epithelial cell of partners, generallyresults marker protein 1) in the modulation of the activity of thebinding partner. Table 1: Gingival Crevicular Fluid (GCF) biomarkers(Table 1 is adapted from Ellias M F et al. “Proteomic Analysis of SalivaIdentifies Potential Biomarkers for Orthodontic Tooth Movement”. TheScientific World Journal. 2012; 2012: 647240. doi: 10.1100/2012/647240.)

Any of the apparatuses and methods described herein may targetbiomarkers present in gingival cervicular fluid (GCF). In general,various molecules are capable of passing through gingival sulcularepithelium and may filter into GCF. Many of these molecules areassociated with remodeling of paradental tissues during situations suchas normal maintenances, periodontal diseases and Orthodontic treatment.The collection and analysis of GCF is a non-invasive procedure that maybe useful information on the nature and extent of the periodontalresponse to mechanotheraphy and orthodontic treatments. The apparatusesand methods described here may be configured and/or adapted to collectGCF. For example any of the apparatuses described herein may include aprojection extending between the teeth and gingiva (e.g., penetrating toa depth of 0.5 mm, 1 mm, 2 mm, 3 mm, 4 mm, 5 mm, 6 mm, etc.). Theprojection may be fixed or extendable. The projection may include acapillary.

Modified or enhanced cellular activities during orthodontic toothmovement can be found in GCF of treated teeth. Additional biomarkersthat may be examined, e.g., in GCF and/or saliva include prostaglandin E(PGE) (elevated prostaglandin E (PGE) levels in GCF 1 day afterapplication of mechanical stimuli has been detected), cytokines,including IL-6 and IL-8, TNF-α, Hyaluronic acid, chondroitin sulphate,IGF, Acid phosphatase, Aspartate aminotransferase, Alkaline phosphatase(ALP), Lactate dehydrogenase, Collagenase, Matrix metalloproteinases(e.g., MMP-1, MMP-2 and MMP-8), Cathepsin B, TRAP, Osteocalcin,Osteonectin, Osteopontin and dentin sialoprotein. For example, acorrelation has been found between the velocity of tooth movement andincrease in concentrations of cytokine and its receptor antagonist.IL-6, IL-8 levels in GCF after force application has been shown.Increased level of TNF-α in GCF after force application which peaked atday 1 has been shown. Studies demonstrated elevated Hyaluronic acid inall GCF samples and chondroitin sulphate levels in GCF increasedgreatest in teeth that moved most. IGF (bone remodeling marker) may beelevated, and its binding protein levels in GCF, 4 h after mechanicalstimulation. Acid phosphatase and Aspartate aminotransferase levelsafter force application are higher on compressed side compared totension side in GCF. Alkaline phosphatase (ALP) levels are higher ontension side compared to compression side. Lactate dehydrogenase levelsare higher on compression side whereas Collagenase levels are elevatedon both mesial and distal sides after mechanical stimulus. Matrixmetalloproteinases (MMPs) (MMP-1, MMP-2 and MMP-8) show elevated levelson compressed side than on tension side. Elevated levels of Cathepsin B,an indicator of ECM degradation, were demonstrated in GCF 1 day afterforce application. Elevated levels of TRAP in GCF on the compressionside after force application has been shown. Osteocalcin, a boneturnover marker, may be elevated in GCF of patients with periodontalbreakdown. Elevated levels of Osteonectin and Osteopontin have beendetected in GCF with progressive increase in periodontal breakdown.Elevated levels of dentin sialoprotein in GCF samples of teeth at 12weeks following commencement of fixed appliance therapy have also beendemonstrated.

For example, the apparatuses and methods described herein may beconfigured to detect one or more of the following biomarkers. In somevariations, these biomarkers may be detected from the saliva and/or thegingival crevicular fluid (GCF). These biomarkers may be particularlyhelpful in detecting tooth remodeling or movement. The levels of one ormore of these biomarkers may be detected and tracked over the course ofa treatment to adjust or modify an orthodontic treatment. For example,one of more markers for inflammation, remodeling and/or enzymes (e.g.,enzymes associated with bone resorption, formation, cell necrosis,collagen remodeling, etc.) may be detected. Examples of makers forremodeling of the teeth may include: Glycosaminoglycans (e.g., in GCF),including hyaluronic acid and a minor band of chondroitin sulfate,Pyridinium derivatives (e.g., pyridinoline and deoxypyridinoline),Pentraxin-3, also known as tumor necrosis factor (TNF)-stimulated gene14 (TSG-14), N-telopeptide type 1 and osteocalcin, Osteocalcin, andMatrix metalloproteins (MMPs) 1 and 8. Markers for inflammation mayinclude: Prostaglandin E (PGE2), Neuropeptides (calcitonin related genepeptide and substance p), Transforming growth factor-α1, Epidermalgrowth factor (EGF), α2 Microglobulin (α2MG), insulin-like growthfactor-1, Interleukin-1 (receptor antagonist) (IL-1) 1β, 2, 6, 8cytokines, Tumor necrosis factor-α, Macrophages colony stimulatingfactors, TNF-related ligand receptor activator of nuclear factor-kappaligand (RANKL) and its two receptors, receptor activator of nuclearfactor-kappa (RANK), and osteoprotegerin (OPG), and Myeloperoxidase(MPO). Markers of root resorption may include: dentine matrix protein 1,dentin phosphoprotein (DPP), and dentin sialoprotein (DSP). Enzymes andenzyme inhibitors may include: Cathepsin B, Acid phosphatase (ACP) andalkaline phosphatase (ALP), β-Glucuronidase (βG), Aspartateaminotransferase (AST), and lactate dehydrogenase.

As mentioned above, any of the biosensors described herein may include abio-recognition component, a biotransducer component, and electronicsystem which may include a signal amplifier, processor, data loggingunits and data communication unit. In some instances, transducers andelectronics can be combined such as in CMOS-based microsensor systems.The recognition component may be called a bioreceptor, may use abiomolecule from organisms or receptors modeled after biological systemsto interact with the analyte of interest. This interaction may bemeasured by the biotransducer which outputs a measurable signalproportional to the presence of the target analyte in the sample. Theprocessor can log the raw or processed data in the memory unit ortransmit it to a receiver. The system can work actively if energizedwith battery, super-capacitor, or an energy harvesting unit, or it mayperform passively upon being energized via induction using an externaldevice, such as cell phone.

In any of the biosensors described herein, the bioreceptor may beconfigured to interact with the specific analyte of interest to producean effect measurable by the transducer. The bioreceptor may have a highselectivity for the analyte among a matrix of other chemical orbiological components. While the type of biomolecule used may varywidely, biosensors may be classified according to common types ofbioreceptor interactions involving, e.g., interactions such as:antibody/antigen, enzymes/ligands, nucleic acids/DNA, cellularstructures/cells, or biomimetic materials. The bioreceptor may beconfigured to engage in one or more of these interactions (e.g., mayinclude a bound or engineered antibody, enzyme, nucleic acid sequence,protein or engineered protein, etc.) in a localized manner that may beinterrogated by or communicated to the biotransducer.

For example, a biosensor as described herein for use with an oralappliance may be configured to take advantage of an antibody/antigeninteraction with one or more of the biomarkers described herein. Thus,the biosensor may be configured as an immunosensor. An immunosensor mayutilize the very specific binding affinity of antibodies for a specificcompound or antigen. The specific nature of the antibody antigeninteraction is analogous to a lock and key fit in that the antigen willonly bind to the antibody if it has the correct conformation (properselection of primary and secondary antibodies). Binding events result ina physicochemical change that, in combination with a tracer, such asfluorescent molecules or enzymes, can generate a signal such as anelevation in voltage that can be detected by electronic components.Binding may be detected optically (e.g., by color, or transmission)and/or electrically.

Also described herein are biosensors that include enzymaticinteractions. For example, analyte recognition may be enabled through:(1) an enzyme converting the analyte into a product that issensor-detectable, (2) detecting enzyme inhibition or activation by theanalyte, or (3) monitoring modification of enzyme properties resultingfrom interaction with the analyte. Since enzymes are not consumed inreactions, the biosensor may be used continuously. The catalyticactivity of enzymes may also lower limits of detection compared tocommon binding techniques.

Other biosensing techniques that can be used may include detectingnucleic acid interactions, detecting epigenetic modifications, cellbased, and tissue based detection.

As mentioned above, a biotransducer may be electrochemical, optical,electronic, piezoelectric, gravimetric, and/or pyroelectric-based.

FIGS. 5A and 5B illustrate an example of an apparatus including abiosensor as described herein. For example, in FIG. 5A, the apparatus500 includes an orthodontic device, shown as an aligner 504 to which abiosensor or biosensors are coupled, either integrated into the housingof the electronics sub-assembly 502, or separate 551 but electricallyconnected to the electronics. The electronics (processor, battery,communications circuitry, antenna, etc.) is shown in a housing thatmounted to an outside portion of the aligner, shown on the buccal side,although it may be on the lingual side instead or additionally. FIG. 5Bshows an example of a section through an example of the apparatus inwhich the biosensor (or a separate sensor 510, such as a temperaturesensor or the like) is included with the electronics 502. An outer cover508 may hold the assembly to the aligner 504 and the apparatus may beplaced over the teeth 506. In some variations the electronics and/orsensor are on the inner surface of the aligner (e.g., the cavity intowhich the teeth sit.

In addition to the biosensors described above, also described herein aresensors configured to determine stress-induced bioelectric potentials onthe teeth. Stress-induced bioelectric potentials may regulate alveolarbone remodeling during orthodontic tooth movement. For example, theforce, F, applied to the labial surface of the lower incisor thatdisplaces the tooth in its socket, deforming the alveolar bone convexlytowards the root at the leading edge, may produce concavity towards theroot at the trailing edge. Concave bone surfaces characterized byosteoblastic activity are electronegative; convex bone surfacescharacterized by osteoclastic activity are electropositive orelectrically neutral. Measuring the electrical charge on the teethsurface, may be used to determine the tooth movement rate and direction.Such data can be used to conduct a closed-loop orthodontic treatment.For example, any of the methods an apparatuses described herein maymeasure or detect the electrical charges from the surface of differentregions of a subject's teeth. An apparatus may include a plurality ofelectrodes within the concavity of an aligner, or a plurality ofelectrical contacts for contacting electrodes attached to the teeth,e.g., attached to one or both of the buccal and lingual sides of any ofthe teeth that are being moved by the orthodontic appliance. Theapparatus may include the electrical system that is configured toreceive these surface charge readings and may store, transmit (e.g.,wireless) or analyze these signals, e.g., in the electrical system orremotely, to determine and/or evaluate forces on the teeth and toothmovement.

Also described herein are systems and apparatuses for determining one ormore indicator of the patient's heath. For example, any of these methodsand apparatuses may include a sensor configured as a physiologicalsensor to detect one or more subject physiological state. For example,any of the apparatuses or methods described herein may measure (e.g.,using a physiological sensor) one or more of: electrocardiogram (ECG),bio-impedance, blood oxygenation, galvanic skin response, heart rate,body temperature, respiration (including respiration rate), or the like.For example, any of these apparatuses may include an ECG sensor (e.g.,one-point ECG electrode), a thermistor, a bio-impedance sensor, aphotoplethysmogram sensor, a galvanic skin response sensor, etc.,including electronics to support such sensor(s). These sensors can beutilized by themselves or with any other sensor, including one or moreof the biosensors described herein. As with any of the biosensors andsensors described herein, these sensors may be used to detect ordetermine compliance (e.g., use of the aligners) while they generatehealth information of the patient. For instance, a photoplethysmogram(PPG) sensor may measure blood-volume changes in the blood tissue. Aplethysmogram is volumetric measurement of an organ. This technique isnon-invasive and may be obtained by illuminating light into the body andmeasuring the change in light absorption. In the current invention, thistechnique may be applied within the intraoral cavity. A plethysmographysensor can be incorporated in an orthodontic appliance (e.g., aligner)to determine the blood volume change in a cheek or within an extendedgingiva segment near or under the appliance to detect blood volumechange in gingiva.

Alternatively or additionally, a galvanic skin response (GSR) sensor maybe used to measure conductivity of intraoral tissues. Conductivity maychange with both changes in the underlying amount of minerals releasedonto the outer surface of tissues from glands.

Also described herein are methods and apparatuses for detecting and/oranalyzing breath. For example any of the apparatuses described hereinmay be used to detect and/or diagnose disease. For instance, lung andbreath cancers may be detected via analysis of the breath, e.g., byidentifying particular breath volatile organic compounds (B VOCs) thatdiffer between patients with non-small cell lung cancer (NSCLC) andsubjects without the disease. Other sensors that detect cancer at earlystages via, for instance, measuring chemical components of breath duringexhale. Exhaled breath contains both volatile and non-volatile organiccompounds, which vary between healthy individuals and those with lungcancer.

The apparatuses and methods described herein may also be configured todetect halitosis (bad breath). Halitosis may arise from inside the mouthand/or due to a disorders in the nose, sinuses, throat, lungs,esophagus, or stomach. Bad breath may also be due to an underlyingmedical condition such as liver failure or ketoacidosis. Halitosis mayalso arise from an underlying disease such as gum disease, tooth decay,or gastroesophageal reflux disease.

By far the most common causes of halitosis are odor producing biofilm onthe back of the tongue, below the gum line, and in the pockets createdby gum disease between teeth and the gums. This biofilm results in theproduction of high levels of foul odors produced mainly due to thebreakdown of proteins into individual amino acids, followed by thefurther breakdown of certain amino acids to produce detectable gases.Volatile sulfur compounds are associated with oral malodor levels, andusually decrease following successful treatment. The intensity of badbreath may differ during the day, due to eating certain foods (such asgarlic, onions, meat, fish, and cheese), smoking, and alcoholconsumption. The odor may be worse upon awakening and may be transientor persistent (e.g., chronic bad breath).

The apparatuses and methods described herein may be configured to detectone or more compounds or markers for bad breath (e.g., above a targetthreshold) that may indicate bad breath, and may alert the wearer,track, store, and/or transmit detected levels. Markers that may bedetected by the apparatuses described herein may include indole,skatole, polyamines, volatile sulfur compounds (VSCs) such as hydrogensulfide, methyl mercaptan, allyl methyl sulfide, and dimethyl sulfide.In some variations the apparatus may detect one or more bacterialmarkers arising due to halitosis-producing bacteria. Bacteria that causegingivitis and periodontal disease (periodontopathogens) may be gramnegative may produce VSC. Methyl mercaptan is known to be a contributingVSC in halitosis that and may be caused by periodontal disease andgingivitis.

These apparatuses may also be used to determine, detect and/or diagnoseother dental issued, including gum disease. For example, the level ofVSC on breath has been shown to positively correlate with the depth ofperiodontal pocketing, the number of pockets, and whether the pocketsbleed when examined with a dental probe. VSC may themselves contributeto the inflammation and tissue damage that is characteristic ofperiodontal disease. Markers for halitosis may also suggest or indicateinfection (e.g., oral infection), oral ulceration, stress/anxiety,menstrual cycle (e.g., at mid cycle and during menstruation, increasedbreath VSC has been reported), or the like. Any of the methods andapparatuses described herein may also or alternatively be used to detector determine (and/or aid in treatment) of any of these indications.

For example, a dental apparatus including a sensor may be configured todetect sulfide (e.g., sulfur emissions) from the patient's breath,saliva and/or GCF. For example, the sensor may be configured to detecthydrogen sulfide. Alternatively or additionally, the apparatus may beconfigured to detect methyl mercaptan, and dimethyl sulfide. In somevariations the sensor may be configured to detect a salivary levels ofan enzyme indicating the presence of certain halitosis-related bacteria,such as β-galactosidase.

Any of the apparatuses described herein may include microfluidics (e.g.,lab-on-a-chip) components. Microfluidics may be used as part of thebiosensor, for example, including channels for acquiring a biologicalfluid (e.g., saliva and/or GCF), processing the fluid (e.g., combiningwith one or more reagents and/or detecting an interaction with abiomolecule, etc.).

Also included herein is the use of one or more bio sensors (e.g.,integrated with an orthodontic appliance such as an aligner) to detecteither a protein or DNA component of an allergen. See, e.g., Alves etal., describing a biosensor system for food allergen detection (e.g.,Alves et al. 2015. DOI: 10.1080/10408398.2013.831026). As describedherein, biosensors are well-suited to automation and their ability todetect multiple analytes in one test with minimal sample preparation andmay be used to conduct in vivo detection and detect the presence of foodallergens in close to real-time. For example, surface plasmon resonance(SPR)-based biosensors, typically used to the fast detection ofegg-related fining allergens in wines, can be integrated with any of theorthodontic appliances (e.g., aligners) described herein to allow rapiddetection the presence of egg white allergens at concentrations between0.03 and 0.20 μg/mL.

Any of the methods (including user interfaces) described herein may beimplemented as software, hardware or firmware, and may be described as anon-transitory computer-readable storage medium storing a set ofinstructions capable of being executed by a processor (e.g., computer,tablet, smartphone, etc.), that when executed by the processor causesthe processor to control perform any of the steps, including but notlimited to: displaying, communicating with the user, analyzing,modifying parameters (including timing, frequency, intensity, etc.),determining, alerting, or the like.

In general, the methods and apparatuses described herein may be used formonitoring the progress of appliance-based orthodontic treatment and/orcompliance. Generally, a monitoring apparatus may include one or morebiosensors and/or sensors (e.g., physiological sensors) configured togenerate sensor data; this data may be related to repositioning of apatient's teeth using an orthodontic appliance. The biosensor and/orsensor data can be processed and analyzed to determine whether theappliance is successfully repositioning the teeth according toprescribed treatment plan. Advantageously, also described herein areintegrated electronic sensing and logging systems capable of generatingmore reliable and accurate aligner performance data, which may be usedby the treating practitioner to track treatment progress and adjust thepatient's treatment plan if desired. The monitoring devices of thepresent disclosure can provide high value sensing data useful foradaptive closed-loop treatment planning and appliance design.

Monitoring performance of an orthodontic appliance for repositioning apatient's teeth is described. The apparatus can comprise an orthodonticappliance comprising one or more teeth-receiving cavities shaped toreposition the patient's teeth from an initial arrangement towards atarget arrangement. The device can comprise one or more sensorsconfigured to generate sensor data related to the repositioning of thepatient's teeth by the orthodontic appliance. The device can comprise aprocessor configured to process the biosensor/sensor data in order toevaluate the performance of the orthodontic appliance in effecting therepositioning of the patient's teeth.

The performance of the orthodontic appliance can be measured in avariety of ways. For example, the processor may be configured toevaluate the performance of the orthodontic appliance by using thebiosensor and/or sensor data to determine one or more of: an amount offorce or pressure applied to the patient's teeth, a distribution offorce or pressure on the patient's teeth, an amount of movement of thepatient's teeth, or a movement rate of the patient's teeth, and/or thephase of movement of the patient's teeth (e.g., initial phase, a lagphase, and a post-lag phase, etc.).

The performance of the orthodontic appliance can determine one or moreof: determining if movement is happening, what phase of tooth movementthe patient is experiencing, and/or the rate of tooth movement. Thisinformation may be processed on the orthodontic appliance itself or offof the appliance (in a remote processor, etc.) and communicated to thedental professional and/or patient. This information may be used toadjust the treatment plan, including instruction removal of anorthodontic appliance (e.g., a removable orthodontic appliance/device)ahead of a scheduled removal, leaving the orthodontic appliance onlonger than a scheduled removal date, or maintaining the scheduledremoval/replacement date. In some variations the information (e.g., theinformation derived by monitoring the level of one or more biomarkersusing the biosensors) may be used to modify the treatment plan bytriggering replacement of one or more devices (aligners) within aplanned sequence of removable orthodontic appliances.

In addition to the biosensors described herein, any of these methods andapparatuses may include a force or pressure sensor configured to measureforce or pressure applied to one or more teeth by the orthodonticappliance. A force or pressure sensor can comprise a force- orpressure-sensitive film, a resistive film, a capacitive film, or apiezoelectric tactile sensor. The processor can be configured toevaluate the performance of the orthodontic appliance by determiningwhether an amount of force or pressure applied to the patient's teeth bythe orthodontic appliance is within a targeted range. Any of thesesensors may be used in combination with one or more biosensor, e.g., toconfirm or estimate movement of the teeth.

A movement sensor may be included and configured to measure movement ofone or more teeth. A movement sensor can comprise an electromagneticfield generator configured to generate an electromagnetic field. Amovement sensor can be configured to measure the movement of the one ormore teeth by measuring changes to the electromagnetic field. Forinstance, a movement sensor can comprise one or more electromagnetictargets arranged to move in response to the movement of the one or moreteeth, such that movement of the one or more electromagnetic targetsproduces changes to the electromagnetic field.

Any of the apparatuses described herein may include a plurality ofdifferent biosensor and/or sensors operably coupled to differentportions of the orthodontic appliance. Any or all of thesebiosensors/sensors can be integrated with the orthodontic appliance,coupled to a tooth, or a combination thereof. As discussed above, aprocessor may be integrated with the orthodontic appliance or coupled toa tooth. Alternatively, a processor can be located external to thepatient's intraoral cavity. Any of these apparatuses my furthercomprises a communication module configured to transmit one or more ofthe sensor data or the processed sensor data to a remote device. In somevariations, the voltage generated by the biosensor may be used torecharge the battery.

A method for monitoring performance of an orthodontic appliance forrepositioning a patient's teeth may include receiving biosensor and/orsensor data related to the repositioning of the patient's teeth by theorthodontic appliance from one or more biosensors and/or sensors. Theorthodontic appliance can comprise one or more teeth-receiving cavitiesshaped to reposition the patient's teeth from an initial arrangementtowards a target arrangement. The biosensor data can be processed inorder to evaluate the performance of the orthodontic appliance ineffecting the repositioning of the patient's teeth.

Performance of the orthodontic appliance may be evaluated by using thebiosensor data to determine one or more of: the state of a biomarkerassociated with tooth movement and/or remodeling, an amount of force orpressure applied to the patient's teeth, a distribution of force orpressure on the patient's teeth, an amount of movement of the patient'steeth, or a movement rate of the patient's teeth.

The apparatuses and methods described herein may include transmitting(e.g., wirelessly transmitting) one or more of the sensor and/orbiosensor data or the processed sensor data to a remote device.

As mentioned above, the methods and apparatuses described herein can beused in combination with various types of orthodontic appliances. Forexample, appliances may have teeth-receiving cavities that receiveand/or reposition teeth, e.g., via application of force due to applianceresiliency, are illustrated in FIGS. 1A and 1B-1D. FIG. 1A illustratesan exemplary tooth repositioning appliance or aligner 100 that can beworn by a patient in order to achieve an incremental repositioning ofindividual teeth 102 in the jaw. The appliance can include a shellhaving teeth-receiving cavities that receive and resiliently repositionthe teeth. An appliance or portion(s) thereof may be indirectlyfabricated using a physical model of teeth. For example, an appliance(e.g., polymeric appliance) can be formed using a physical model ofteeth and a sheet of suitable layers of polymeric material. In someembodiments, a physical appliance is directly fabricated, e.g., usingrapid prototyping fabrication techniques, from a digital model of anappliance.

The methods and apparatuses described herein may be used with anyappliance that receives teeth, for example appliances without one ormore of polymers or shells. The appliance can be fabricated with one ormore of many materials such as metal, glass, reinforced fibers, carbonfiber, composites, reinforced composites, aluminum, biologicalmaterials, and combinations thereof for example. The appliance can beshaped in many ways, such as with thermoforming or direct fabrication(e.g., 3D printing, additive manufacturing), for example. Alternativelyor in combination, the appliance can be fabricated with machining suchas an appliance fabricated from a block of material with computernumeric control machining.

An appliance can fit over all teeth present in an upper or lower jaw, orless than all of the teeth. The appliance can be designed specificallyto accommodate the teeth of the patient (e.g., the topography of thetooth-receiving cavities matches the topography of the patient's teeth),and may be fabricated based on positive or negative models of thepatient's teeth generated by impression, scanning, and the like.Alternatively, the appliance can be a generic appliance configured toreceive the teeth, but not necessarily shaped to match the topography ofthe patient's teeth. In some cases, only certain teeth received by anappliance will be repositioned by the appliance while other teeth canprovide a base or anchor region for holding the appliance in place as itapplies force against the tooth or teeth targeted for repositioning. Insome embodiments, some, most, or even all of the teeth will berepositioned at some point during treatment. Teeth that are moved canalso serve as a base or anchor for holding the appliance as it is wornby the patient. Typically, no wires or other means will be provided forholding an appliance in place over the teeth. In some cases, however, itmay be desirable or necessary to provide individual attachments or otheranchoring elements 104 on teeth 102 with corresponding receptacles orapertures 106 in the appliance 100 so that the appliance can apply aselected force on the tooth.

As used herein, a dental appliance may include an aligner, such as thoseutilized in the Invisalign® System, which are described in numerouspatents and patent applications assigned to Align Technology, Inc.,including, for example, in U.S. Pat. Nos. 6,450,807, and 5,975,893, aswell as on the company's website, which is accessible on the World WideWeb (see, e.g., the url “invisalign.com”). In this specification, theuse of the terms “orthodontic aligner”, “aligner”, or “dental aligner”may be synonymous with the use of the terms “appliance” and “dentalappliance” in terms of dental applications. For purposes of clarity,embodiments are hereinafter described within the context of the use andapplication of appliances, and more specifically “dental appliances.”

Examples of tooth-mounted attachments suitable for use with orthodonticappliances are also described in patents and patent applicationsassigned to Align Technology, Inc., including, for example, U.S. Pat.Nos. 6,309,215 and 6,830,450.

FIGS. 1B-1D illustrate a tooth repositioning system 110 including aplurality of appliances 112, 114, 116. Any of the appliances describedherein can be designed and/or provided as part of a set of a pluralityof appliances used in a tooth repositioning system. Each appliance maybe configured so a tooth-receiving cavity has a geometry correspondingto an intermediate or final tooth arrangement intended for theappliance. The patient's teeth can be progressively repositioned from aninitial tooth arrangement to a target tooth arrangement by placing aseries of incremental position adjustment appliances over the patient'steeth. For example, the tooth repositioning system 110 can include afirst appliance 112 corresponding to an initial tooth arrangement, oneor more intermediate appliances 114 corresponding to one or moreintermediate arrangements, and a final appliance 116 corresponding to atarget arrangement. A target tooth arrangement can be a planned finaltooth arrangement selected for the patient's teeth at the end of allplanned orthodontic treatment. Alternatively, a target arrangement canbe one of some intermediate arrangements for the patient's teeth duringthe course of orthodontic treatment, which may include various differenttreatment scenarios, including, but not limited to, instances wheresurgery is recommended, where interproximal reduction (IPR) isappropriate, where a progress check is scheduled, where anchor placementis best, where palatal expansion is desirable, where restorativedentistry is involved (e.g., inlays, onlays, crowns, bridges, implants,veneers, and the like), etc. As such, it is understood that a targettooth arrangement can be any planned resulting arrangement for thepatient's teeth that follows one or more incremental repositioningstages. Likewise, an initial tooth arrangement can be any initialarrangement for the patient's teeth that is followed by one or moreincremental repositioning stages.

The orthodontic appliances described herein can be fabricated in a widevariety of ways. As an example, some embodiments of the appliancesherein (or portions thereof) can be produced using indirect fabricationtechniques, such as by thermoforming over a positive or negative mold.Indirect fabrication of an orthodontic appliance can involve producing apositive or negative mold of the patient's dentition in a targetarrangement (e.g., by rapid prototyping, milling, etc.) andthermoforming one or more sheets of material over the mold in order togenerate an appliance shell. Alternatively or in combination, someembodiments of the appliances herein may be directly fabricated, e.g.,using rapid prototyping, stereolithography, 3D printing, and the like.

The configuration of the orthodontic appliances herein can be determinedaccording to a treatment plan for a patient, e.g., a treatment planinvolving successive administration of a plurality of appliances forincrementally repositioning teeth. Computer-based treatment planningand/or appliance manufacturing methods can be used in order tofacilitate the design and fabrication of appliances. For instance, oneor more of the appliance components described herein can be digitallydesigned and fabricated with the aid of computer-controlledmanufacturing devices (e.g., computer numerical control (CNC) milling,computer-controlled rapid prototyping such as 3D printing, etc.). Thecomputer-based methods presented herein can improve the accuracy,flexibility, and convenience of appliance fabrication.

Orthodontic appliances, such as the appliance illustrated in FIG. 1A,may impart forces to the crown of a tooth and/or an attachmentpositioned on the tooth at one or more points of contact between a toothreceiving cavity of the appliance and received tooth and/or attachment.The magnitude of each of these forces and/or their distribution on thesurface of the tooth can determine the type of orthodontic toothmovement which results. Tooth movements may be in any direction in anyplane of space, and may comprise one or more of rotation or translationalong one or more axes. Types of tooth movements include extrusion,intrusion, rotation, tipping, translation, and root movement, andcombinations thereof, as discussed further herein. Tooth movement of thecrown greater than the movement of the root can be referred to astipping. Equivalent movement of the crown and root can be referred to astranslation. Movement of the root greater than the crown can be referredto as root movement.

FIG. 2 illustrates a method 200 of orthodontic treatment using aplurality of appliances, in accordance with embodiments. The method 200can be practiced using any of the appliances or appliance sets describedherein. In step 210, a first orthodontic appliance is applied to apatient's teeth in order to reposition the teeth from a first tootharrangement to a second tooth arrangement. In step 220, a secondorthodontic appliance is applied to the patient's teeth in order toreposition the teeth from the second tooth arrangement to a third tootharrangement. The method 200 can be repeated as necessary using anysuitable number and combination of sequential appliances in order toincrementally reposition the patient's teeth from an initial arrangementto a target arrangement. The appliances can be generated all at the samestage or time point, in sets or batches (e.g., at the beginning of oneor more stages of the treatment), or one at a time, and the patient canwear each appliance until the pressure of each appliance on the teethcan no longer be felt or until the maximum amount of expressed toothmovement for that given stage has been achieved. A plurality ofdifferent appliances (e.g., a set) can be designed and even fabricatedprior to the patient wearing any appliance of the plurality. Afterwearing an appliance for an appropriate period of time, the patient canreplace the current appliance with the next appliance in the seriesuntil no more appliances remain. The appliances are generally notaffixed to the teeth and the patient may place and replace theappliances at any time during the procedure (e.g., patient-removableappliances). The final appliance or several appliances in the series mayhave a geometry or geometries selected to overcorrect the tootharrangement. For instance, one or more appliances may have a geometrythat would (if fully achieved) move individual teeth beyond the tootharrangement that has been selected as the “final.” Such over-correctionmay be desirable in order to offset potential relapse after therepositioning method has been terminated (e.g., permit movement ofindividual teeth back toward their pre-corrected positions).Over-correction may also be beneficial to speed the rate of correction(e.g., an appliance with a geometry that is positioned beyond a desiredintermediate or final position may shift the individual teeth toward theposition at a greater rate). In such cases, the use of an appliance canbe terminated before the teeth reach the positions defined by theappliance. Furthermore, over-correction may be deliberately applied inorder to compensate for any inaccuracies or limitations of theappliance.

FIG. 2B illustrates a method 1100 for monitoring performance of anorthodontic appliance for repositioning a patient's teeth using abiosensor and/or sensor. The method 1100 can be performed using anyembodiment of the apparatuses described herein. Some or all of the stepsmay be performed using a processor of a monitoring device operablycoupled to an orthodontic appliance. Alternatively or in combination,some or all of the steps can be performed by a processor of a deviceexternal to the patient's intraoral cavity, e.g., a separate computingdevice or system. As used herein, a monitoring device may refer to theelectronics system (subsystem) and may optionally include the one ormore biosensors and/or sensors, and may include a housing or case,covering the electronics, power supply, etc.

In step 1110, biosensor and/or sensor data is received from one or morebiosensors (and/or sensors) operably coupled to an orthodonticappliance. The one or more biosensors can include any of the biosensortypes described herein, including biosensors testing saliva and/or GCF.

The orthodontic appliance can be worn by the patient as part of atreatment plan for incrementally repositioning the patient's teeth. Theorthodontic appliance may include teeth-receiving cavities shaped toreposition one or more teeth according to a prescribed treatment plan,and the biosensor(s) can be physically integrated with (e.g., coupledto, embedded in, formed with, etc.) the orthodontic appliance atlocations adjacent to or near the teeth to be repositioned. Thebiosensor data can be related to the repositioning of the patient'steeth by the orthodontic appliance. For example, the biosensor data canprovide information regarding movements (e.g., rotational,translational) of one or more teeth. As another example, the biosensordata can provide information regarding the interaction between theorthodontic appliance and the patient's teeth or attachment devicesmounted thereto. The biosensor data may be generated and loggedcontinuously. Alternatively, in order to reduce power consumption, thebiosensor data can be obtained at predetermined time intervals, such asonce every 15 minutes, 30 minutes, 1 hour, 2 hours, 5 hours, 12 hours,or 24 hours. The timing for biosensor and/or sensor data collection mayvary based on the expected tooth movements to be produced by theorthodontic appliance. For example, in some embodiments, tooth tippingis expected to occur relatively rapidly after the patient starts wearingthe appliance, such that monitoring for tooth tipping is performedduring the first 12 hours of appliance usage.

In step 1120, the biosensor and/or sensor data is processed in order toevaluate the performance of the orthodontic appliance in repositioningthe patient's teeth. For example, the biosensor data can includeconfirmation that the appliance has triggered remodeling of teeth withina predetermined amount, and the processing step can involve determiningwhether the analyte indicating remodeling is within a targeted range ofvalues, e.g., for repositioning the teeth. Alternatively or incombination, the biosensor and/or sensor data can include measurement ofchanges in the spatial disposition (e.g., position and/or orientation)of one or more teeth, and the processing step can involve determiningwhether the changes in spatial disposition correspond to plannedmovements for the patient's teeth. Optionally, the processing step caninvolve associating the biosensor data with a timestamp representingwhen the data was obtained such that appliance performance informationcan be measured over time.

The processed biosensor data can include appliance performanceinformation, e.g., whether the force(s), pressure(s), and/or toothmovement(s) produced by the appliance correlate well with the expectedvalues for the planned orthodontic treatment. The expected values for aplanned treatment may be determined by computer simulation. For example,an orthodontic appliance can be considered to be performingsatisfactorily if: (1) the measured force and/or pressure values liewithin the expected range for those values, or is within 70% of atargeted value; (2) the pattern of force and/or pressure application onthe teeth matches, or is similar to, the planned pattern for forceand/or pressure application; (3) the amount of tooth movement achievedis within 70% of the planned movement; (4) the direction of toothmovement matches, or is similar to, the planned direction of toothmovement; or combinations thereof. An orthodontic appliance can beconsidered to be performing unsatisfactorily if: (1) the measured forceand/or pressure values lie outside the expected range for those valuesor is more than 30% away from a targeted value; (2) the pattern of forceand/or pressure application on the teeth differs from the plannedpattern for force and/or pressure application; (3) the amount of toothmovement achieved is more than 30% away from the planned movement; (4)the direction of tooth movement is different to the planned direction oftooth movement; or combinations thereof.

In step 1130, the biosensor data generated in step 1110 and/or theprocessed biosensor data generated in step 1120 are optionallytransmitted to a remote device. The remote device can be a mobile device(e.g., smartphone), personal computer, laptop, tablet, wearable device,cloud computing server, or the like. Step 1130 can be performed usingwireless or wired communication methods, as desired. Step 1130 can beperformed automatically (e.g., at predetermined time intervals) or inresponse to instructions received from the remote device (e.g., acommand to transmit the biosensor and/or sensor data and/or applianceusage).

In step 1140, the orthodontic treatment plan prescribed to the patientis optionally modified based on the biosensor and/or sensor datagenerated in step 1110 and/or the processed biosensor data generated instep 1120. The modification step can be performed by a processorexternal to the patient's intraoral cavity, such as a remote device asin step 1130. Modifying the treatment plan can involve modifying aplanned intermediate or final arrangement of the patient's teeth,modifying the teeth-receiving cavity geometries of an orthodonticappliance corresponding to a planned intermediate or final tootharrangement, modifying the timing for wearing one or more appliances,modifying the order for wearing a series of appliances, or a combinationthereof. For example, if the appliance performance information indicatesthat the tooth repositioning achieved by the orthodontic appliance isnot satisfactory and the teeth are off-track, the treatment plan can bemodified in order to move the patient's teeth back on track (e.g.,mid-course correction). As another example, if the appliance performanceinformation indicates that the appliance is not producing the desiredforce and/or pressure pattern on the teeth, the geometries of subsequentappliances can be adjusted accordingly to provide more accurate forceand/or pressure application. By using the appliance performanceinformation as feedback, these methods and apparatuses allow foradaptive, closed-loop orthodontic treatment based on the actual responseof the patient's teeth to treatment.

The monitoring devices described herein can be physically integratedinto an orthodontic appliance in a variety of ways. In some embodiments,the monitoring device is integrated into the appliance during or afterfabrication of the appliance. For example, the monitoring device can beattached to an appliance using adhesives, fasteners, a latchingmechanism, or a combination thereof after the appliance has beenfabricated. Optionally, the appliance can be formed with complementaryfeatures or structures (e.g., recesses, receptacles, guides, apertures,etc.) shaped to receive and accommodate the monitoring device orcomponents thereof.

A monitoring device may be coupled to the appliance as a prefabricatedunit during or after fabrication of the appliance, such as by beinginserted and sealed into a receptacle in the appliance, attached to anappliance (e.g., by a latching mechanism, adhesive, fastener).Alternatively, the monitoring device can be assembled in situ on theappliance during or after appliance fabrication. For instance, inembodiments where the appliance is manufactured by direct fabrication(e.g., 3D printing), the monitoring device can be printed simultaneouslywith the appliance, inserted into the appliance during fabrication, orafter assembled the appliance has been fabricated. Optionally, some ofthe monitoring device components may be prefabricated and othercomponents may be assembled in situ. It shall be appreciated that thevarious fabrication methods described herein can be combined in variousways in order to produce an appliance with integrated monitoring devicecomponents.

An orthodontic appliance can be operably coupled to a monitoring deviceconfigured to provide data related to tooth repositioning and/or theinteraction between the appliance and the patient's teeth (e.g., contactbetween the appliance and the teeth, the amount of force and/or pressureapplied by the appliance to the teeth, distribution of force and/orpressure on the teeth, etc.). Such data can be used to evaluate theperformance of the orthodontic appliance for repositioning the patient'steeth. For instance, appliance performance information as describedherein can include information regarding whether the force(s),pressure(s), and/or tooth movement(s) produced by an orthodonticappliance correlate with the expected values for the planned orthodontictreatment.

The monitoring devices described herein can be designed for use in thepatient's intraoral cavity. For example, the dimensions of a monitoringdevice may be limited in order to avoid patient discomfort and/orfacilitate integration into an orthodontic appliance as discussed below.In some embodiments, a monitoring device has a height or thickness lessthan or equal to about 1.5 mm, or less than or equal to about 2 mm. Insome embodiments, a monitoring device has a length or width less than orequal to about 4 mm, or less than or equal to about 5 mm. The shape ofthe monitoring device can be varied as desired, e.g., circular,ellipsoidal, triangular, square, rectangular, etc. For instance, in someembodiments, a monitoring device can have a circular shape with adiameter less than or equal to about 5 mm.

A relatively thin and flexible monitoring device can be used to providea larger surface area while reducing patient discomfort. In someembodiments, the monitoring devices herein are sized to conform to asurface of a tooth crown (e.g., a buccal, lingual, and/or occlusalsurface of a tooth crown). For example, a monitoring device havingdimensions of about 10 mm by about 5 mm can be used to cover a buccalsurface of a molar crown. As another example, a monitoring device havingdimensions of about 10 mm by about 20 mm can be used to cover thebuccal, occlusal, and lingual surfaces of a tooth crown. A monitoringdevice can be in contact with a crown of a single tooth, or with crownsof a plurality of teeth, as desired.

The monitoring device dimensions (e.g., volume, weight) can be designedin order to reduce patient discomfort. For instance, the weight of amonitoring device can be selected not to exceed a level that would exertundesirable forces on the underlying teeth. A monitoring device may beused primarily for research and characterization purposes, rather thanfor patient treatment, and thus may not be subject to size constraintsfor reducing patient discomfort. For example, in embodiments where themonitoring device is used outside the intraoral cavity (e.g., benchtoptesting of aligner performance), the size of the monitoring device canbe relatively large compared to devices designed for intraoral use.

As discussed above, FIG. 3 schematically illustrates an example of anapparatus (e.g., a monitoring device 300 portion of an apparatus), thatmay be used or include an orthodontic appliance (not shown). Themonitoring device 300 can be used in combination with any embodiment ofthe systems and devices described herein, and the components of themonitoring device 300 are equally applicable to any other embodiment ofthe apparatuses, including monitoring devices, described herein. Themonitoring device 300 can be implemented as an application-specificintegrated circuit (ASIC) including one or more of the followingcomponents: a processor 302, a memory 304, one or more biosensors and/orsensors 306, a clock 308, a communication unit 310, an antenna 312, apower management unit 314, or a power source 316. The processor 302(e.g., a central processing unit (CPU), microprocessor, fieldprogrammable gate array (FPGA), logic or state machine circuit, etc.),also referred to herein as a controller, can be configured to performthe various methods described herein. The memory 304 encompasses varioustypes of memory known to those of skill in the art, such as RAM (e.g.,SRAM, DRAM), ROM (EPROM, PROM, MROM), or hybrid memory (e.g., flash,NVRAM, EEPROM), and the like. The memory 304 can be used to storeinstructions executable by the processor 302 to perform the methodsprovided herein. Additionally, the memory can be used to storebiosensor/sensor data obtained by the biosensor(s)/sensors 306, asdiscussed in greater detail below.

The monitoring device 300 can include any number of biosensors 306and/or sensor 306′, such as one, two, three, four, five, or morebiosensors. In some embodiments, the use of multiple biosensors providesredundancy to increase the accuracy and reliability of the resultantdata. Some or all of the biosensors 306 can be of the same type. Some orall of the biosensors 306 can be of different types. Examples ofbiosensor types suitable for use in the monitoring devices describedherein are provided below. Examples of additional sensors may include:touch or tactile sensors (e.g., capacitive, resistive), proximitysensors, movement sensors (e.g., electromagnetic field sensors), forcesensors (e.g., force-sensitive resistive or capacitive materials),pressure sensors (e.g., pressure-sensitive resistive or capacitivematerials), strain gauges (e.g., resistive- or MEMS-based), electricalsensors, or combinations thereof.

A biosensor 306 can be operably coupled to and/or located at any portionof an orthodontic appliance, such as at or near a distal portion, amesial portion, a buccal portion, a lingual portion, a gingival portion,an occlusal portion, or a combination thereof. A biosensor 306 can bepositioned near a tissue of interest when the appliance is worn in thepatient's mouth, such as near or adjacent the teeth, gingiva, palate,lips, tongue, cheeks, airway, or a combination thereof. For example,when the appliance is worn, the biosensor(s) 306 can cover a singletooth, or a portion of a single tooth. Alternatively, the biosensor(s)306 can cover multiple teeth or portions thereof. In embodiments wheremultiple biosensors 306 are used, some or all of the monitoring devicescan be located at different portions of the appliance and/or intraoralcavity. Alternatively, some or all of the biosensors 306 can be locatedat the same portion of the appliance and/or intraoral cavity.

An analog-to-digital converter (ADC) (not shown) can be used to convertanalog biosensor and/or sensor data into digital format, if desired. Theprocessor 302 can process the data obtained by the biosensor(s) 306 inorder to determine appliance usage and/or patient compliance, asdescribed herein. The biosensor data and/or processing results can bestored in the memory 304. Optionally, the stored data can be associatedwith a timestamp generated by the clock 308 (e.g., a real-time clock orcounter).

In some embodiments, the monitoring device 300 includes a communicationunit 310 configured to transmit the data stored in the memory (e.g.,biosensor data and/or processing results) to a remote device. Thecommunication unit 310 can utilize any suitable communication method,such as wired or wireless communication methods (e.g., RFID, near-fieldcommunication, Bluetooth, ZigBee, infrared, etc.). The communicationunit 310 can include a transmitter for transmitting data to the remotedevice and an antenna 312. Optionally, the communication unit 310includes a receiver for receiving data from the remote device. In someembodiments, the communication channel utilized by the communicationunit 310 can also be used to power the device 300, e.g., during datatransfer or if the device 300 is used passively.

The remote device can be any computing device or system, such as amobile device (e.g., smartphone), personal computer, laptop, tablet,wearable device, etc. Optionally, the remote device can be a part of orconnected to a cloud computing system (“in the cloud”). The remotedevice can be associated with the patient, the treating practitioner,medical practitioners, researchers, etc. In some embodiments, the remotedevice is configured to process and analyze the data from the monitoringdevice 300, e.g., in order to assess appliance performance, for researchpurposes, and the like.

The monitoring device 300 can be powered by a power source 316, such asa battery. In some embodiments, the power source 316 is a printed and/orflexible battery, such as a zinc-carbon flexible battery, azinc-manganese dioxide printed flexible battery, or a solid-state thinfilm lithium phosphorus oxynitride battery. The use of printed and/orflexible batteries can be advantageous for reducing the overall size ofthe monitoring device 300 and avoiding patient discomfort. For example,printed batteries can be fabricated in a wide variety of shapes and canbe stacked to make three-dimensional structures, e.g., to conform theappliance and/or teeth geometries. Likewise, flexible batteries can beshaped to lie flush with the surfaces of the appliance and/or teeth.Alternatively or in combination, other types of batteries can be used,such as supercapacitors. In some embodiments, the power source 316 canutilize lower power energy harvesting methods (e.g., thermodynamic,electrodynamic, piezoelectric) in order to generate power for themonitoring device 300. Optionally, the power source 316 can berechargeable, for example, using via inductive or wireless methods. Insome embodiments, the patient can recharge the power source 316 when theappliance is not use. For example, the patient can remove theorthodontic appliance when brushing the teeth and place the appliance onan inductive power hub to recharge the power source 316.

Optionally, the apparatus can include a power management unit 314connected to the power source 316. The power management unit 314 can beconfigured to control when the apparatus is active (e.g., using powerfrom the power source 316) and when the apparatus inactive (e.g., notusing power from the power source 316). In some embodiments, themonitoring device 300 is only active during certain times so as to lowerpower consumption and reduce the size of the power source 316, thusallowing for a smaller monitoring device 300

The apparatus may also include an activation mechanism (not shown) forcontrolling when the monitoring device (e.g., control circuitry) 300 isactive (e.g., powered on, monitoring appliance usage) and when themonitoring device 300 is dormant (e.g., powered off, not monitoringappliance usage). The activation mechanism can be provided as a discretecomponent of the monitoring device 300, or can be implemented by theprocessor 302, the power management unit 314, or a combination thereof.The activation mechanism can be used to reduce the amount of power usedby the monitoring device 300, e.g., by inactivating the device 300 whennot in use, which can be beneficial for reducing the size of the powersupply 316 and thus the overall device size.

In some embodiments, the monitoring device 300 is dormant before beingdelivered to the patient (e.g., during storage, shipment, etc.) and isactivated only when ready for use. This approach can be beneficial inconserving power expenditure. For example, the components of themonitoring device 300 can be electrically coupled to the power source316 at assembly, but may be in a dormant state until activated, e.g., byan external device such as a mobile device, personal computer, laptop,tablet, wearable device, power hub etc. The external device can transmita signal to the monitoring device 300 that causes the activationmechanism to activate the monitoring device 300. As another example, theactivation mechanism can include a switch (e.g., mechanical, electronic,optical, magnetic, etc.), such that the power source 316 is notelectrically coupled to the other components of the monitoring device300 until the switch is triggered. For example, the switch may be a reedswitch or other magnetic sensor that is held open by a magnet. Themagnet can be removably attached to the monitoring device 300, or may beintegrated into the packaging for the device 300 or appliance, forexample. When the monitoring device is separated from the magnet (e.g.,by removing the magnet or removing the device and appliance from thepackaging), the switch closes and connects the power source 316, asillustrate in FIG. 4B. As another example, the monitoring device 300 caninclude a mechanical switch such as a push button that is manuallyactuated in order to connect the power source 316. In some embodiments,the activation mechanism includes a latching function that locks theswitch upon the first actuation to maintain connectivity with the powersource so as to maintain activation of the monitoring device 300.Optionally, the switch for the activation mechanism can be activated bya component in the patient's intraoral cavity (e.g., a magnet coupled toa patient's tooth), such that the monitoring device 300 is active onlywhen the appliance is worn by the patient, and is inactive when theappliance is removed from the patient's mouth. Alternatively or incombination, the switch can be activated by other types of signals, suchas an optical signal.

FIG. 4B illustrates a monitoring device 400 with an activationmechanism. The monitoring device 400, as with all other monitoringdevices described herein, can be similar to the monitoring device 300,and can include some or all of the components described herein withrespect to the monitoring device 300. The device 400 is coupled to anorthodontic appliance 402 (e.g., via an encapsulating material 404). Thedevice 400 can include an activation mechanism 403 including a magneticswitch. Prior to use, the device 400 can be removably coupled to amagnet 406 (e.g., using tape 408), and the magnet 406 can hold themagnetic switch in an open position such that the device 400 isinactive. When the appliance 402 is ready for use, the user can removethe magnet 406, thus closing the magnetic switch and connecting thecomponents of the monitoring device 400 to a power source.

The orthodontic appliances and monitoring devices can be configured inmany different ways. In some embodiments, an orthodontic appliance maybe operably coupled to a single monitoring device. Alternatively, theorthodontic appliance can be operably coupled to a plurality ofmonitoring devices, such as at least two, three, four, five, or moremonitoring devices. Some or all of the monitoring devices may be of thesame type (e.g., collect the same type of data). Alternatively, some orall of the monitoring devices may be of different types (e.g., collectdifferent types of data). Any of the embodiments of monitoring devicesdescribed herein can be used in combination with other embodiments in asingle orthodontic appliance.

A monitoring device can be located at any portion of the appliance, suchas at or near a distal portion, a mesial portion, a buccal portion, alingual portion, a gingival portion, an occlusal portion, or acombination thereof. The monitoring device can be positioned near atissue of interest when the appliance is worn in the patient's mouth,such as near or adjacent the teeth, gingiva, palate, lips, tongue,cheeks, airway, or a combination thereof. For example, when theappliance is worn, the monitoring device can cover a single tooth, or aportion of a single tooth. Alternatively, the monitoring device cancover multiple teeth or portions thereof. In embodiments where multiplemonitoring devices are used, some or all of the monitoring devices canbe located at different portions of the appliance. Alternatively, someor all of the monitoring devices can be located at the same portion ofthe appliance.

A monitoring device can be operably coupled to the orthodontic appliancein a variety of ways. For example, the monitoring device can bephysically integrated with the orthodontic appliance by coupling themonitoring device to a portion of the appliance (e.g., using adhesives,fasteners, latching, laminating, molding, etc.). The coupling may be areleasable coupling allowing for removal of the monitoring device fromthe appliance, or may be a permanent coupling in which the monitoringdevice is permanently affixed to the appliance. Alternatively or incombination, the monitoring device can be physically integrated with theorthodontic appliance by encapsulating, embedding, printing, orotherwise forming the monitoring device with the appliance. In someembodiments, the appliance includes a shell shaped to receive thepatient's teeth, and the monitoring device is physically integrated withthe shell. The monitoring device can be located on an inner surface ofthe shell (e.g., the surface adjacent to the received teeth), an outersurface of the shell (e.g., the surface away from the received teeth),or within a wall of the shell. Optionally, as discussed further herein,the shell can include a receptacle shaped to receive the monitoringdevice. Exemplary methods for fabricating an appliance with a physicallyintegrated monitoring device (e.g., by incorporating some or all of thecomponents of the monitoring device during direct fabrication of theappliance) are described in further detail herein.

FIGS. 5A and 5B illustrate an example of an apparatus including anorthodontic appliance 500 having an integrated monitoring device(control circuitry) 502 and biosensor. In this example, the appliance500 includes a shell 504 having one or more (e.g., a plurality of)teeth-receiving cavities, and the monitoring device 502 is coupled to anouter, buccal surface of the shell 504 adjacent a tooth receiving cavity506. In the depicted embodiment, the monitoring device 502 is coupled toa tooth receiving cavity 506 for a molar. It shall be appreciated thatin alternative embodiments, the monitoring device 502 can be coupled toother portions of the shell 504, such as an inner surface, a lingualsurface, an occlusal surface, one or more tooth receiving cavities forother types of teeth (e.g., incisor, canine, premolar), etc. Themonitoring device 502 can be shaped to conform to the geometry of thecorresponding appliance portion (e.g., the wall of the cavity 306) so asto provide a lower surface profile and reduce patient discomfort. Insome embodiments, the appliance 500 includes a receptacle 508 formed onthe outer surface of the shell 504 and the monitoring device 502 ispositioned within the receptacle. Exemplary methods for forming anappliance with a receptacle 508 and integrated monitoring device 502 aredescribed in detail below.

The monitoring device 502 can include any of the components previouslydescribed herein with respect to the monitoring device 300 of FIG. 3.For example, the monitoring device 502 can include a biosensor 510and/or sensor, a power source 512 (e.g., a battery), and/or acommunication unit 514 (e.g., a wireless antenna). The arrangement ofthe components of the monitoring device 502 can be varied as desired. Insome embodiments, the biosensor 551 is located adjacent to the toothreceiving cavity 506. A gap can be formed in the shell 504 adjacent tothe biosensor/sensor 510 so as to permit direct access to the receivedtooth. The communication unit 514 (or a component thereof, such as anantenna) can be located adjacent to or on the outer surface of thereceptacle 408 so as to facilitate data transmission.

Some of the components of a monitoring device may be packaged andprovided separately from other components of the device. For example, amonitoring device can include one or more components that are physicallyintegrated with a first orthodontic appliance and one or more componentsthat are physically integrated with a second orthodontic appliance. Thefirst and second orthodontic appliances can be worn on opposing jaws,for example. Any of the components of a monitoring device (e.g.,components of the device 300 of FIG. 3) can be located on an appliancefor the upper jaw, an appliance for the lower jaw, or a combinationthereof. In some embodiments, it is beneficial to distribute thecomponents of the monitoring device across multiple appliances in orderto accommodate space limitations, accommodate power limitations, and/orimprove sensing, for example. Additionally, some of the components of amonitoring device can serve as a substrate for other components (e.g., abattery serves as a substrate to an antenna).

Alternatively or in combination, other types of biosensor/sensor can beused to indirectly measure the forces and/or pressures applied to theteeth by an appliance. For example, in some embodiments, the applicationof force and/or pressure to a patient's teeth produces electricalcurrents (for example, via the piezoelectric effect) in structures ofthe mouth. Compression of bone and collagen may result in movement ofelectrons in the crystal lattice, and application of force on the teethcan result in a short piezoelectric effect on the alveolar bone, whichmay be detected by appropriate receiving sensors such as electrodes.Electrical signals produced by alveolar and periodontal ligaments (PDL)when under load can stimulate changes in bone metabolism. Thispiezoelectric effect can be measured to determine when a tooth is loadedor overloaded by an appliance. Electrical sensors such as electrodes mayalso be used to detect these electrical signals, for example, bymonitoring changes in voltage.

Alternatively or in combination, the monitoring devices herein caninclude one or more tactile sensors that respond to direct contact withthe patient's teeth. The tactile sensors described herein can becapacitive sensors, resistive sensors, inductive sensors, orpiezoelectric sensors, for example. For example, the tactile sensor canbe a piezoelectric sensor including one or more materials that exhibitpiezoelectric properties, such as quartz, ceramics, or polymers (e.g.,polyvinylidene fluoride (PVDF)).

In some embodiments, a biosensor can be a sensor array that capable ofdetecting contact over a two-dimensional surface area. Optionally, atactile sensor can be provided as a clear, thermoformable screen or filmcapable of conforming to the shape of the appliance. Some types oftactile sensors may only be capable of providing contact data (e.g.,binary data indicating the presence or absence of direct contact), whileother types of tactile sensors may also be capable of providing othertypes of data in addition to contact data (e.g., resistive tactilesensors capable of providing force and/or pressure data).

A monitoring device can include a single biosensor, or a plurality ofbiosensors and/or other sensors can be positioned at any location in theappliance, such on an inner surface, an outer surface, a buccal surface,a lingual surface, an occlusal surface, a mesial portion, a distalportion, a gingival portion, or a combination thereof. In embodimentswhere the orthodontic appliance includes a shell with a teeth-receivingcavity, the biosensors/sensors can be positioned on the inner surfacesof the teeth-receiving cavities. Optionally, at least some biosensorscan be located on an outer surface of the appliance, such as an occlusalsurface in order to detect contact between the upper and lower teeth

The biosensors can be positioned to be near certain teeth when theappliance is worn, e.g., near teeth to be repositioned and/or atlocations where the appliance is expected to exert force on the teeth.For example, tactile sensors can be located at or near the buccal,lingual, and/or occlusal surfaces of a tooth to be repositioned so as toprovide a map of contact points over the tooth crown. In someembodiments, the monitoring device is configured to obtain data frombuccal, lingual, and occlusal sensors in a predetermined order and at adesired frequency in order to provide a contact map over the buccal,lingual, and occlusal surfaces. Alternatively or in combination, if theappliance is shaped to engage an attachment device mounted on a tooth, atactile sensor can be located at or near the location of engagementbetween the appliance and the attachment device.

Alternatively or in combination, an apparatus as described herein caninclude one or more movement sensors for measuring the movements (e.g.,translational and/or rotational movements) of one or more teeth. Forexample, a movement sensor can be used track the movements of one ormore teeth relative to the underlying jaw (e.g., mandible or maxilla).As another example, a movement sensor can be used to track the movementsof a first set of one or more teeth relative to a second set of one ormore teeth, such as when tracking the movements of opposite sides of asingle arch during arch or palate expansion. Optionally, a movementsensor can be used to track movements of the upper and lower archesrelative to each other, such as when correcting the relative positioningof the arches in order to treat overbite or underbite. Various types ofmovement sensors can be used. In some embodiments, a movement sensorincludes an electromagnetic field generator (e.g., an electromagneticcoil, generator antenna) integrated into an orthodontic appliance or anattachment device mounted on a patient's tooth. The generator can beconfigured to generate an electromagnetic field (e.g., electric field,magnetic field, or combination thereof) within the intraoral cavity. Themovement sensor can also include one or more electromagnetic targets(e.g., a cylindrical or flat coil, magnet, etc.) integrated into anorthodontic appliance (e.g., the same appliance as the generator, adifferent appliance worn on the opposite jaw, or a combination thereof).The electromagnetic targets can be positioned in the appliance at ornear locations where tooth movement is expected to occur (e.g., coupledto teeth-receiving cavities of teeth to be repositioned), such that themovements of the teeth produce corresponding movements of theelectromagnetic targets. Alternatively or in combination, the monitoringdevice can include one or more electromagnetic targets integrated intoan attachment device coupled to the patient's teeth, such that themovements of the teeth and associated targets are directly correlated.

Alternatively or in combination, any of the apparatuses described hereincan include one or more electrical sensors (e.g., electrodes) to measuretooth surface charges, as mentioned above. Alveolar bone remodelingduring orthodontic tooth movement may be regulated by stress-inducedbioelectric potentials on the tooth surface. For example, a forceapplied to the labial surface of the lower incisor can displace thetooth in its socket, deforming the alveolar bone convexly towards theroot at the leading edge, and producing concavity towards the root atthe trailing edge. In some embodiments, concave bone surfacescharacterized by osteoblastic activity are electronegative, and convexbone surfaces characterized by osteoclastic activity are electropositiveor electrically neutral. Accordingly, the monitoring device can measurethe electrical charges on the tooth surface in order to determine thetooth movement rate and/or direction.

Alternatively or in combination, any of the apparatuses described hereincan include one or more conductivity sensors configured to measure theconductivity of fluids (e.g., saliva) in the surrounding environment. Insome embodiments, bone remodeling during orthodontic tooth movementcauses changes in saliva content, and these changes can be measuredbased on the ionic charge of the minerals in the saliva. Examples ofminerals that may influence the conductivity of saliva include but arenot limited to NH4+, Ca2+, P043″, HC03-, and F″.

In general, the apparatuses described herein may include miniaturizedand integrated electronic components (e.g., battery, antenna,controller, wireless communication circuitry, etc.) as part of anembedded biosensing apparatus. In some variations, the biosensor mayinclude detection of one or more types of biomarker in saliva, includingthose in Table 1, above. For example, a Potentiostat with ascreen-printed electrode and a modified enzyme layer may be used todetect a salivary biomarker (e.g., 14-3-3 protein σ (Stratifin), uricacid, etc.). A working electrode (biotransducer) may be chemicallymodified by crosslinking to an enzyme. An antifouling layer may beincluded to prevent interference effects and biofouling.

The apparatus may include electrodes for differential C2D (e.g., commonto differential) measurements and input channels for A2D (analog todigital) voltage measurements. One or more electrodes may be layeredwith an enzyme-membrane to achieve different configurations of workingelectrodes. Additional sensors and/or biosensors may be used, includingtemperature measurements. For example, one or more sensors can provideproximity data, which can augment biomarker detection.

In any of these apparatuses and methods described herein, compliancedata may be determined from the biosensor/sensor data, and thisinformation may also be used to augment, control, and/or interpret thebiosensor information. For example, compliance data may be estimated bydetermining a working state of the apparatus from proximity data (e.g.,power states for “In-mouth” and “out-of-mouth” conditions). Temperaturesensing may also be used to augment biomarker data, e.g., by correlatingtemperature and biomarker data, which may provide more specificphysiological monitoring.

As discussed above, examples of biomarkers that may be detected by theapparatuses and methods herein may include salivary biomarkers such assRANKL, OPG, which may be correlated to different phases of orthodontictooth movement (e.g., https://www.ncbi.nlm.nih.gov/pubmed/23273364).Other salivary markers include S100-A9, immunoglobulin J chain, Igalpha-1 chain C region, CRISP-3, which may indicate inflammation andbone resorption (see, e.g.,https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3417200/). Examples ofgingival crevicular fluid biomarkers, e.g., inflammatory fluidaccessible in in the gingival margin, may include prostaglandin E2(which may indicate bone resorption), Substance P (neuropeptide) (whichmay indicate bone resorption), epidermal growth factor (which mayindicate bone resorption), transforming growth factor (which mayindicate bone remodeling), RankL (which may indicate stimulation ofosteoclsatic differentiation), Granulocyet macrophage colony stimulationfactor (which may indicate bone turnover), a2 microglobulin-enhance ofIGF 1, Interleukin 1β, 2, 6, 8 (which may indicate bone remodeling),Myeloperoxidase (and enzyme involved in PMN inflammation). Othergingival crevicular fluid biomarkers may include glycosaminoglycans(GAGs or mucopolysaccharides), and may indicate paradental remodeling,such as hyaluronic acid (a type of GAG or mucopolysaccharide, indicatorof breakdown of gingival tissue), and Chondroitin sulfate (another typeof GAG, an indicator of breakdown of alveolar bone and PDL).

Also described herein are textile-based sensors. Such sensors mayinclude stretchable conductive ink, and may be used when forming theapparatus (e.g., on an aligner). For example, a textile-based sensor maybe co-formed with the apparatus or added to it, or added directly ontothe teeth. In some variations the sensor including a conductive “ink”may be screen printed as a trace on a flexible substrate, includingfabric substrates or polymeric substrates. The appliance (e.g., aligner)may then be formed specific to the patient, as described above. Theflexible substrate with the printed sensor and/or trace may then beapplied (or alternatively screened directly) on/over the the aligner.Thus, the stretchable conductive ink may be directly applied over thealigner.

Alternatively, as mentioned, a sensor, such as a stretchable sensor, maybe bonded directly to the teeth or other intra oral tissue. This mayprovide better access to GCF or saliva for the biosensor/sensor. Insteadof an embedded biosensor on the aligner, the biosensor may be directlybonded to the gingival margin to monitor GCF for certain biomarkers.

Any of the biosensors/sensors described herein may be powered by aflexible fuel cell, such as a stretchable rechargeable battery. Thus,the entire sensing apparatus or sub-system may be flexible and/or may bebonded directly to the tooth. Alternatively or additionally, a sensor(broadly including a biosensor) may be powered and/or augmented byelectronics in an appliance (e.g., aligner). For example, the sensor mayneed to be bonded to the tooth for better monitoring of thephysiological signal, but driving electronics may be located on areplaceable aligner (for renewed energy supply, data collection, signalprocessing, etc.).

FIGS. 6A-6C illustrate an example of an apparatus bonded directly to thesubject's teeth. In FIG. 6A, the apparatus includes a stretchableconductive trace that is directly attached to the teeth. In thisexample, the traces connect nodes A and B to the power supply nodes Cand D, which are on the aligner. This configuration may eliminateleakage current from the battery to the components (X) when the aligneris not in the mouth. In one example, the components generically referredto as “X” in FIG. 6A-6C could include electronics which drive a BLEsignal every few minutes (e.g., every minute, 2 minutes, 3 minutes, 5minutes, 10 minutes, 15 minutes, etc., or variable intervals/times). Areceiver (e.g., smartphone and/or dedicated receiver) could track theBLE pulses and monitor when the appliance is in the mouth.

In FIG. 6A, the traces 603 shown on the teeth 601 can be used to connectto rigid components on the aligner. Different electronic circuits may beused when the aligner is in the mouth and for when the aligner isoutside of the mouth. For example, traces such as those shown above canbe used as performance measures for aligners or tooth movements. Nodes607 can be placed to be at known positions on the aligner and comparedwith nodes positions on teeth. In some variations a printedpotentiometer may be applied to the teeth. As shown in FIG. 6B, beforethe aligner is worn, the contacts (A, B) or nodes are not connected, tothe power (on left, nodes D and C), which may be on the aligner. Whenthe aligner is worn (shown in FIG. 6C), the circuit is completed, asnodes A and D connect and nodes B and C connect by the conductive trace.At a minimum, this completed circuit may be used to indicate compliance,as it will only complete the circuit when the appliance is worn, andworn correctly. Alternatively, additional sensor(s), including one ormore biosensor, may also be connected to the traces on the subject'steeth (either on the tooth/teeth, or on the aligner) and activated whenthe appliance is worn. In some variations, the use of conductive traceson the teeth that may interface with contacts on an aligner may also beused to check the fit of an aligner.

In some variations the traces, such as those shown in FIGS. 6A-6C may bemagnetic, which may allow self-healing of the traces, or may also beused for other purposes, including detection (e.g., via a reed switch orhall-effect sensor, etc.) including detection of teeth.

As mentioned above, any of the biosensor systems and apparatuses (e.g.,removable orthodontic devices) described herein may be used to monitorone or more biomarkers from a patient. For example, FIG. 8 schematicallyillustrates one example of a method of monitoring a biomarker from apatient using a removable orthodontic device, such as an aligner, thatincludes a biomarker. As mentioned above, as an initial step (not shown)a removable orthodontic device that does not apply substantial force tomove the teeth but that includes a biosensor system may be first worn,and a baseline for the one or more biomarkers collected. In FIG. 8, aremovable orthodontic device (e.g., aligner) having a biosensor may beworn by the patient. For example, the removable orthodontic device mayhave a plurality of tooth receiving cavities that may be placed incommunication with a patient's teeth (e.g., one or more of the patient'steeth). The plurality of tooth receiving cavities may be configured toexert one or more orthodontic repositioning forces on the patient'steeth 802.

While the aligner is worn, the bioreceptor (that may be housed in abiosensor housing) of the removable orthodontic aligner may be placed incontact with a fluid (e.g., saliva, GCF, blood) within the oral cavity,which may cause a first interaction with one or more biomarkers fortooth motion (“tooth motion biomarkers”). The first interaction may berelated to a change in expression of a first biomarker of the one ormore tooth motion biomarkers, wherein the change in expression of thefirst biomarker is associated with a specific phase of tooth movement ofone or more of the patient's teeth 804. For example, the level of thebiomarker compared to a baseline may be indicative of the phase of toothmovement (e.g., initial phase, lag phase, etc.). A threshold or range ofsensed values may be used to monitor an effect of the removableorthodontic device on the teeth. For example, the first interaction maybe transduced into a first interaction signal representative of thefirst interaction 806, and this first interaction signal may be providedby the device 808. For example it may be output (transmitted, displayed,stored, etc.).

FIG. 9 illustrates an example of a method of modifying an orthodontictreatment plan using a removable orthodontic device including abiosensor. In FIG. 9, the biosensor signal may be received by aprocessor on the removable orthodontic device and/or by a remoteprocessor (e.g. a smartphone, computer, server, etc.). For example, themethod may include receiving a first interaction signal representing afirst interaction between the bioreceptor in a removable orthodonticaligner and one or more tooth motion biomarkers in fluid (e.g., saliva,GCF, blood, etc.) in a fluid in the patient's oral cavity, the removableorthodontic aligner configured to receive a plurality of a patient'steeth and to exert one or more orthodontic repositioning forces on theplurality of the patient's teeth 902. The interaction signal may beanalog or digital, and may be stored and/or processed (filtered,amplified, etc.) including normalized to another biomarker signal, acontrol signal and/or a baseline.

The first biomarker expression change of the tooth motion biomarker maybe identified 904. This first biomarker expression change may be relatedto the first interaction between the bioreceptor and the one or moretooth motion biomarkers. For example, one or more specific phase(s) oftooth movement of one or more teeth in the oral cavity of the patientmay be identified using the first biomarker expression change 906.

The method may also include determining whether to gather one or morerecommendations to modify an orthodontic treatment plan implementing theremovable orthodontic device based on the specific phase of toothmovement 908. For example, a recommendation may be based on the level ofthe one or more biomarker signals, as described above. Thisrecommendation may be derived from stored logic, and/or a memory (e.g.,a look-up table, by machine learning, etc.).

Microfluidics

Any of the biosensors described herein may include one or moremicrofluidics systems for capture, storage and analysis of intra-oralfluids, including chemical analysis. For example, any of thesemicrofluidic systems may include hard or flexible/stretchable (such assilicone) materials with or without integrated electronics (includingwireless communication electronics), and may be formed integrally withthe apparatus (e.g., aligner) and/or be intimately and robustly bond tothe surface of apparatus.

In some variations, the microfluidic system may include a network ofreservoirs for embedded chemical agents that may respond in colorimetricfashion to biomarkers. The reservoirs may be connected by microfluidicschannels. In some variations the microfluidics channels may beconfigured for active and/or passive metering, so that a fluid fromwithin the patient's oral cavity (e.g., saliva and/or GCF) may be drawninto the microfluidics channel and passed into a sample chamber. Thesample chamber may include, for example a colorometric indicator orother chemical agent that responds to one or more biomarkers in thefluid in a colorimetric manner. Alternatively, in some variations, thesample is processed in a microfluidics channel for later read-out (e.g.,when removing the device from the mouth, and placing it into a separatestorage and/or readout chamber.

In any of these variations, apparatus may include microfluidic channelsthat are configured to allow access to various sample and/or detectionregions on the apparatus at various times. For example, themicrofluidics device integrated into or on an aligner may be configuredto provide timing via chrono-sampling of a fluid. For example, amicrofluidic system can be designed to enable sampling withchronological order and controlled timing. In some variations, thetiming of fluid within the microchannel may be timed actively, e.g., bythe opening of a channel via release of a valve (e.g., anelectromechanical valve, an electromagnetic value, a pressure valve,etc.). Examples of valves controlling fluid in a microfluidic networkinclude piezoelectric, electrokinetics and chemical approaches.Capillary bursting valves (CBVs) are another variation of a valve for amicrofluidics channel. CBVs block flows at pressures lower than theircharacteristic bursting pressures (BPs). When liquid in a singleconnected channel encounters two separate CBVs with different BPs, atsufficient pressures, the flow will proceed first through the valve withlower BP. In this way, locating two CBVs with different BPs near theintersection between two channels allows control of the direction offlow. The Young-Laplace equation gives the BP in a rectangular channel:

$\begin{matrix}{{BP} = {{- 2}{\sigma \left\lbrack {\frac{\cos \; \theta_{1}^{*}}{b} + \frac{\cos \; \theta_{A}}{h}} \right\rbrack}}} & \lbrack 1\rbrack\end{matrix}$

-   -   where σ is the surface tension of liquid, θ_(A) is the contact        angle of the channel, θ₁* is the min[θ_(A)+β; 180° ], β is the        diverging angle of the channel, b and h are the width and the        height of the diverging section, respectively. For hydrophobic        materials at high diverging angles, the BP increases with        decreasing b and h.

Thus, by adjusting the angles and dimensions, the bursting pressure maybe adjusted and in the context of a microfluidics channel, series ofCBVs may be used to set up a sequence of timed regions that open as thefluid within the channel reaches the selected BP. For example, thediverging angles may be between 13° and 90°, or 13° and 120°. One ofskill in the art would therefore be able to select the microfluidicchannel lengths and BP configurations to arrange a series ofmicrofluidics channels and chambers, which are valved by one or morecontrol valves, including CBVs that open at predetermined time ranges toallow sampling over time.

In some variations the microfluidic channel may be opened by dissolvinga material blocking the channel. The rate of dissolution may becalibrate to open the channel after a predetermined time period (e.g.,minutes, hours, days, etc.). In some variations the dissolving materialmay include one or more reagents for use in detecting and/or storing thefluid in the microfluidics apparatus. For example, a blocking materialmay include a labeling material (e.g., such as an antibody, enzyme,substrate, etc.) for detection within a chamber blocked by the blockingmaterial.

For example, FIGS. 10A-10H illustrate an example of an apparatus,configured as an aligner, that may be worn on a subject's teeth toprovide sampling and/or storage of a fluid from the patient's oralcavity. In FIG. 10A, the aligner 1001 includes one or moretooth-receiving portions 1003 and is configured to be won over thepatient's teeth. In this example, a microfluidics component 1005 mayform part of a biosensor and may be included or incorporated into thealigner, as shown. FIG. 10B shows an enlarged view of the microfluidicsportion of the exemplary apparatus. In this example, the microfluidicsportion (e.g., of the biosensor) includes a plurality of microfluidicschannels 1007 and chambers 1009 that are sequentially arranged withinthe device to “open” at different times; fluid (e.g., saliva, GCF) isdrawn into the microfluidics portion at a metered rate toward eachchamber. In addition, the lengths of microfluidics channels 1007increases toward each chamber, requiring additional time for eachchannel to reach each chamber. The chambers are connected bymicrofluidics channels and the fluid may require a predetermined amountof time to reach each chamber. In some variations the chamber includes aportion of an assay (e.g. a colorimetric indicator) for detecting one ormore biomarker. In FIG. 10B, each microfluidics chamber may also includea vent for preventing air bubbles or build-up in the channel(s); thisvent may be a hydrophobic, air-permeable membrane that permits air toleave the microfluidics channel, but not allow additional saliva to passinto the channel/chamber. In the example shown in FIG. 10B, each channelinto a chamber and from the chamber into the next length of channel(they are arranged in sequence) includes a valve 1013, 1013′. Forexample, the valve(s) may be CBVs as discussed above.

By staggering the timing of opening (access) through the microfluidicchannels into the various chambers, the sampling and/or testing of thefluid from the oral cavity may be controlled. For example, the apparatusof FIGS. 10A-10B may be configured to sample fluid from the saliva everyx hours (e.g., every 1 hour, every 2 hours, every 3 hours, ever 4 hours,every 6 hours, every 12 hours, every 24 hours, every 36 hours, every 48hours, etc.). This is illustrated in FIGS. 10C-10H, showing staggeredfilling of sampling chamber 1021 at approximately 1 hour (FIG. 10C), 12hours (FIG. 10D), 24 hours (FIG. 10E), 36 hours (FIG. 10F), 48 hours(FIG. 10G) and 60 hours (FIG. 10H); at each interval, an additionalsampling/storage chamber is opened.

FIG. 11 is another example of a timed microfluidics sampling and/ortesting region 1031. In this example, six microfluidic chambers 1039 areshown, each connected to a microfluidics channel 1034 that is metered bya separate valve 1032 that may be opened to allow fluid to flow into themicrofluidics channel and into the chamber. The valve may be controlledactively or passively as mentioned above. In some variations thechambers include a preservative material and the material with thechambers 1039 may be held for later testing.

In some variations, one or more multiplexers may be used to sampleand/or measure multiple biomarkers in controlled intervals. For example,a multiplexer may be used to provide access to one or more sampleregions (chambers). The multiplexer and/or any active valves may beconnected and controlled by control circuitry in the apparatus.

Variations

Also described herein are biosensors that are configured to use abiofuel as part of the biosensor. For example an electrochemical sensormay be formed using an enzyme that catalyzes a reaction in the presenceof a biomarker to be identified; the reaction may be detected directly,e.g., when the enzymatic reaction results in an ionizing reaction and iscoupled to an electrical conductor, or indirectly. For example, whenusing biofuel as biosensor, the biomarker may be detected, for example,by measuring how much voltage is generated in the biofuel. Enzymaticbiofuels may work based on ionic current. For example, amperometricelectrodes may be used to detect the level or presence of biomarkers,including ionic species, such as O₂, H₂O₂, NADH, I₂, etc. Similarly,ion-selective electrodes may be used (e.g., to detect pH, NH₄, NH₃, CO₂,I⁻, CN^(−′) etc. Any of these sensors may include a membrane (e.g.,semi-permeable or selectively permeable membrane.

Other examples of biosensors may include paper-based biosensors that maybe integrated with an apparatus such as an aligner. The main constituentof paper is cellulose fiber, and this can be highly attractive forcertain applications, as it allows liquid to penetrate within itshydrophilic fiber matrix without the need of an active pump or externalsource. Moreover, cellulose fibers can be functionalized, thus changingproperties such as hydrophilicity, if desired, as well as itspermeability and reactivity. The paper sensor may be at least partiallyenclosed by the aligner material, and may be directly exposed to theoral cavity or indirectly exposed (e.g., via a channel, including amicrofluidics channel). The paper sensor may be formed of a fibrousmaterial such as nitrocellulose, and may be patterned (e.g., viaphotolithography) or by printing including silk-screening-likeprocesses. The biosensor may include a biomarker that is impregnatedand/or applied and/or absorbed onto the paper into a detection pattern.Multiple positive/negative regions may be created. For example, aglucose-detecting region may use iodine, which may be enzymaticallyreduced in the presence of glucose, resulting in a detectable colorchange (e.g., from clear to brown). Thus, in general, any of thebiosensors described herein may be configured to be used with acellulose (paper) substrate.

In general, as mentioned above, sampling of biosensors can be conductedon an apparatus such as an aligner in an intraoral cavity and thebiochemical or optical assays can be done in the aligner case. Forinstance, saliva or GCF sampling can be conducted using microfluidicsystem or paper-based system while the aligner is worn. When the alignerremoved from the mouth and is placed in the case, optical assessment ofthe concentration of biomarkers in the sampled fluid can be done using avery small optical system integrated into the case. For example, theoptical system may include one or more LEDs, and one or morephotodiodes.

Any of the methods and apparatuses described herein may be used with oneor more biosensors configured for lateral flow and/or vertical flowimmunoassay for biomarker detection. Vertical flow immunoassays rely onthe same basic principles as the more common lateral flow immunoassayformat with some modifications. The most apparent difference between thetwo methods being the vertical and lateral flow of fluid. However,vertical flow technology may have advantages over traditional lateralflow assays, including reduced assay time (<5 minutes).

Vertical flow immunoassays can be used for rapid detection of anantigen(s) including biomarkers. Detection sensitivity may be in thelower nanogram per ml range even in complex sample matrices. Forexample, the biosensor may include an immobilized capture agent (e.g.,antibody) on a reagent pad to which a sample such as saliva or GCF (withor without biomarker to be detected) is applied, e.g., via amicrofluidics channel. Detection of the bound capture agent may beachieved through the binding of, for example, an antigen specificantibody conjugated to a detectable marker, such as a gold conjugate.This step completes a sandwich consisting of a capture agent (e.g.,antibody), the biomarker (antigen) and finally the detectable marker(e.g., gold conjugate) and results in a direct and permanent visuallydetectable marker, such as a red dot indicating the presence of thebiomarker. Alternative colors for detection may be achieved by usingdifferent types of detectable markers, such as nanoparticles conjugatedto a probe for detection.

Biosensors integrated with aligners can be used for detection ofbiomarkers associated with oral and periodontal diseases such as, forexample: DNA probes for detection of putative periodontal pathogens(e.g., Porphyromonas gingivalis, Tanerella forsythensis, Treponemadenticola, Actinobacillus actinomycetemcomitans, etc.), host responsefactors (e.g., IL-1β; TNF-α; aspartate aminotransferase; elastase),connective tissue breakdown products (e.g., collagen telopeptides;osteocalcin; proteoglycans; fibronection fragments, etc.).

Any of the apparatuses including biosensors and methods described hereinmay use aptamers. For example, any of these biosensors may beaptomer-based biosensors. Aptamers are single-stranded nucleic acidsthat selectively bind to target molecules. Aptamers may have highstability, resistance to denaturation and degradation, and may be easilymodified.

EXAMPLES

A biosensor for use with an orthodontic device such as an aligner mayinclude an enzyme or other protein-binding agent (e.g., antibodyfragment, etc.) that is formed into a biosensor by a modifiedscreen-printing electrode formation method. The apparatus may alsoinclude any of the electronics systems described herein, including acontroller (e.g., microcontroller) and wireless communicationsub-system, such as a Bluetooth (e.g., Bluetooth low energy, BLE)transceiver. The biosensor may be formed suing screen-printingtechnology onto a flexible substrate, such as PET (polyethyleneterephthalate). The binding protein (e.g., enzyme) may be crosslinked toa reporter enzyme or indicator that, when the target protein is bound,activates the reporter so that this activity can be detected by thebiotransducer. For example, in some instances the binding protein is anenzyme that engages with the target protein and has as a by-product aperoxide (e.g., H₂O₂); alternatively the binding protein may be modifiedsuch that it is conjugated to a peroxide-forming enzyme and when thetarget biomolecule is present (e.g., in a saliva sample), the enzyme isallowed to reach to form peroxide. In such cases, the bioreporter mayinclude a material, such as Prussian-blue carbon, reacts with thebyproduct to produce a current. For example, in biosensors in whichperoxide is formed in the presence of the target biomarker, a currentmay therefore be detected as hydrogen peroxide contacts the bioreporter(e.g., Prussian Blue carbon). For example, the binding protein may befused with an enzyme (e.g., peroxidase); in the presence of thebiomarker to which the binding protein binds, the enzyme is free toconvert substrate (preferably the biomarker) into an oxidized or reducedform to generate an electrical current on the biosensor, particularlycompared to the reference/control electrode, which may not include thebinding protein.

In one example, a conductive ink may be printed onto a substrate (e.g.,an Ag/AgCl conductive ink) to form the base of an electrode, contactpad/region and one or more traces. A reference electrode as well as abiosensor electrode may be formed; the biosensor may act as anelectrochemical sensor. Interaction and/or binding of the biomarkerprotein to the bioreceptor may be detected either directly by thebioreporter (e.g., as it produces an oxidation or reduction of thebioreporter) or indirectly. Thus, both active (biosensor) and controlelectrodes may include the bioreporter (e.g., graphite ink such asPrussian-blue graphite ink in variations in which a peroxide is formedby the interaction with the biomarker). The bioreporter may also beprinted onto the working (biosensor) and counter electrodes. Aninsulator layer (e.g., of dielectric material) may also be used. In somevariations a sacrificial layer of material that is consumed orconsumable by the bioreceptor and/or bioreporter when interacting withthe biomarker may also be used. After each printing step, the printedlayers may be allowed to cure (e.g., at room temperature or greater).Any appropriate size electrode (working and counter electrodes) may beused, for example, 0.5 mm diameter, 1 mm diameter, 2 mm diameter, 3 mmdiameter, 4 mm diameter, etc. The biosensor may then be modified toinclude the protein that interacts with the biosensor, such as anenzyme, antibody fragment, etc., and an antibiofoulding coating and/ormembrane may be used (e.g., electropolymerized o-phenylenediamine (PPD).

Thus, a biosensor as described herein may be printed by, e.g., ascreening technique in which a substrate (which may be the aligner) issecured and a conductive layer (e.g., Ag/AgCl-based ink) is firstlyapplied to define the conductive underlayer of the biosensor as well asa reference electrode; the conductive layer may be patterned directlyonto the substrate. Next, a catalytic layer may be applied. For example,a carbon or metal-based ink containing any associated catalytic orbiocatalytic functionality may be applied onto the conductor to definethe working and counter electrode. Thereafter, an insulator may beapplied on the conductor and catalytic layers to insulate all but thecontact pads and the upper portion of the electrodes. Subsequent to eachprinting routine, the sensor may be annealed in a temperature-controlledoven at an appropriate temperature to evaporate and volatile solvents.Thereafter, electrochemical activation of the active material may beperformed via repetitive cyclic voltammograms or extended durationamperometry. Cyclic voltammetry may also be executed in a sulfuric acidsolution of moderate strength in order to electrochemically clean theelectrode surface and attenuate any impurities that may interfere withmeasurements. Electrochemical deposition of conducting polymers (i.e.poly(pyrrole), poly(aniline)) or plating of metal catalysts (i.e.palladium, platinum, gold) may also be executed to functionalize/preparethe electrode surface for enhanced detection. Dry reagents,electroactive mediators, and/or permselective membranes may then bedispensed on the surface to achieve pH adjustment, reduce theoverpotential required to excite the electroactive analyte of interest,and/or reject potential interfering compounds. Alternatively, ratherthan screening, these layers may be applied by stamping.

FIGS. 7A-7C illustrate another example of a biosensor that may beincluded as part of a dental apparatus. In this example the biosensordetects a change in pressure due to swelling of a material thatincreases and/or decreases volume with binding to a biomarker. Forexample a sensor may be made based on binding of the biomarker to thegel, which may change the ionic conductivity of the gel, resulting inswelling. The swelling of the gel may be detected mechanically (e.g., bya force sensor), electrictromechanically (e.g., using a microcantilever)and/or optically (e.g., using an optical fiber). In one example, aglucose sensor may be formed by including a glucose-binding agent (e.g.,phenylboronic acid) attached to a gel. A change in ionic conductivity ofthe gel may be measured as the binding reaction changes the swelling ofthe gel.

In some variations the gel may be coated to a force sensor (e.g., agel-coated silicon microcantilever) to and the force sensor may be usedto detect swelling in response to changes in the biomarker species. Forexample, a pH sensors may be formed using a gel that swells/contractsbased on pH. In addition, a sensor to detect enzymatic reactions (enzymeactivity) may be formed by tethering an enzymatic target (or the enzymeitself) to a hydrogel. Enzymatic activity may result induce swelling,which may be detected. In some variations, swelling may be caused by anionic concentration (e.g., Calcium concentration). A crosslinkedpolyacrylic acid gel, which can undergo single-chain aggregation in thepresence of Ca²⁺ may be used to detect local calcium concentration byswelling/shrinking based on the concentration of calcium.

FIG. 7A shows an example of a dental appliance, configured as an aligner704 including a biosensor 700. This biosensor may be permanently orremovably mounted to the outside and/or inside of the appliance by amount 715. A schematic of the biosensor is shown in FIGS. 7B and 7C. Thebiosensor may include a sample port 703 exposing a swellable hydrogel705 that is configured to change volume (e.g., swell) when exposed toone or more particular biomarker. The change in volume may be detectedby the control circuitry 709 within the biosensor. For example thecontrol circuitry may include a force sensor that determines the forcedue to swelling, and/or by electrical changes (e.g., changes inconductivity/resistivity) due to binding and/or activity of the targetbiomolecule. The control circuitry may also include any of the othercomponents described above (e.g., one or more processors, memories,power regulator circuitry, wireless communication circuitry, etc. FIG.7B shows the biosensor prior to swelling and FIG. 7C shows the biosensorafter/during swelling.

Customizing Patient Treatment Using Biomarker Data

In any of the methods and apparatuses described herein, one or morebiomarkers may be monitored and the data from monitoring the biomarkermay be used to modify an orthodontic treatment plan. Monitoring may becontinuous, e.g., using a dental appliance that includes one or moresensors within the patient's oral cavity, as described above, or it maybe performed by sampling from the oral cavity and processing the sampleoutside of the oral cavity.

The dental treatment may be modified base on the absence or presence ofa maker and/or the level (e.g., a normalized level) of one or moremarkers. For example, a patient-specific orthodontic treatment plan maybe modified based on a biomarker detected during a treatment stage. Inone variation a marker for one or more of inflammation (e.g., acuteinflammation, such as cytokines and/or prostaglandins) and/or boneremodeling (e.g., Glycosaminoglycans) may be detected following aninitial first or second treatment stage. In some variations, one or moreinitial measurements, e.g., baseline, may be used specific to a patientand compared to later measurements to detect a change. If the change inlevel of one or more biomarkers is below a minimum threshold (e.g., nochange, change below about x %, where x is 5%, 10%, 15%, 20%, 25%, 30%,35%, 40%, 50%, 60%, 70%, 80%, 90%, etc. compared to a baseline and/orstandard value), then the orthodontic treatment plan may be modified toincrease the amount of movement during the next stage. If the change inlevel of one or more biomarkers is above a maximum threshold (e.g.,change of greater than y %, where y is 30%, 40%, 50%, 60%, 70%, 80%,90%, 100%, 110%, 120%, 130%, 140%, 150%, 160%, 170%, 180%, 190% 200%,etc.) then the orthodontic treatment plan may be modified to decreasethe relative tooth movement during the next stage and/or the duration ofthe current stage.

Alternatively, or additionally, the biomarkers level(s) may be used toadjust the duration that one or more dental appliances (e.g., aligners)in a series of progressive dental appliances are worn. For example, ifthe one or more biomarkers consistent with an acute stage of toothmovement remain elevated during a treatment stage, the duration of thatstage may be extended.

Thus, a series or sequence of dental appliances may be modified based onthe level(s) of one or more biomarkers, including modifying the dentalappliances (e.g., adjusting tooth movement at each stage of the seriesor sequence) and/or adjusting the duration that the one or moreappliances is worn (e.g., wearing them for longer or shorter timeperiods), and/or wearing them for a lesser or greater part of the day.In some variations, the duration of that a final appliance (e.g., aretainer) is to be worn may be determined at least in part by monitoringone or more biomarkers, such as markers of bone remodeling, markers forinflammation, etc. For example, a retainer may be worn until one or morebiomarkers returns to within some predetermined range of a baselineand/or standard value. For example, a patient may continue to wear aretainer until the levels of one or more biomarkers (e.g.,Calgranulin-B, Serum albumin precursor, Immunoglobulin J chain, Igalpha-1 chain C region, Cysteine-rich secretory protein 3 precursor(CRISP-3), Hemoglobin subunit beta, Stratifin, and soluble RANK Ligand(sRANKL), prostaglandin E2, Substance P, epidermal growth factor,transforming growth factor, Receptor activator of nuclear factor kappa-Bligand (RANKL), Granulocyet macrophage colony stimulation factor, α2microglobulin, Interleukin 1β, Myeloperoxidase, hyaluronic acid and/orChondroitin sulfate, etc.) is within a predetermined range of thebaseline and/or standard value (e.g., within about 5%, 10%, 15%, 20%,25%, 30%, 35%, 40%, 50%, etc. of the baseline and/or a standard value).

As discussed above, in some variations, an orthodontic treatment planmay be modified or customized based on biosensor data. For example, aseries of aligners may include one or more aligners with one or moresensor for detecting a biomarker while wearing the apparatus. One ormore sensors may record data.

When a feature or element is herein referred to as being “on” anotherfeature or element, it can be directly on the other feature or elementor intervening features and/or elements may also be present. Incontrast, when a feature or element is referred to as being “directlyon” another feature or element, there are no intervening features orelements present. It will also be understood that, when a feature orelement is referred to as being “connected”, “attached” or “coupled” toanother feature or element, it can be directly connected, attached orcoupled to the other feature or element or intervening features orelements may be present. In contrast, when a feature or element isreferred to as being “directly connected”, “directly attached” or“directly coupled” to another feature or element, there are nointervening features or elements present. Although described or shownwith respect to one embodiment, the features and elements so describedor shown can apply to other embodiments. It will also be appreciated bythose of skill in the art that references to a structure or feature thatis disposed “adjacent” another feature may have portions that overlap orunderlie the adjacent feature.

Terminology used herein is for the purpose of describing particularembodiments only and is not intended to be limiting of the invention.For example, as used herein, the singular forms “a”, “an” and “the” areintended to include the plural forms as well, unless the context clearlyindicates otherwise. It will be further understood that the terms“comprises” and/or “comprising,” when used in this specification,specify the presence of stated features, steps, operations, elements,and/or components, but do not preclude the presence or addition of oneor more other features, steps, operations, elements, components, and/orgroups thereof. As used herein, the term “and/or” includes any and allcombinations of one or more of the associated listed items and may beabbreviated as “/”.

Spatially relative terms, such as “under”, “below”, “lower”, “over”,“upper” and the like, may be used herein for ease of description todescribe one element or feature's relationship to another element(s) orfeature(s) as illustrated in the figures. It will be understood that thespatially relative terms are intended to encompass differentorientations of the device in use or operation in addition to theorientation depicted in the figures. For example, if a device in thefigures is inverted, elements described as “under” or “beneath” otherelements or features would then be oriented “over” the other elements orfeatures. Thus, the exemplary term “under” can encompass both anorientation of over and under. The device may be otherwise oriented(rotated 90 degrees or at other orientations) and the spatially relativedescriptors used herein interpreted accordingly. Similarly, the terms“upwardly”, “downwardly”, “vertical”, “horizontal” and the like are usedherein for the purpose of explanation only unless specifically indicatedotherwise.

Although the terms “first” and “second” may be used herein to describevarious features/elements (including steps), these features/elementsshould not be limited by these terms, unless the context indicatesotherwise. These terms may be used to distinguish one feature/elementfrom another feature/element. Thus, a first feature/element discussedbelow could be termed a second feature/element, and similarly, a secondfeature/element discussed below could be termed a first feature/elementwithout departing from the teachings of the present invention.

Throughout this specification and the claims which follow, unless thecontext requires otherwise, the word “comprise”, and variations such as“comprises” and “comprising” means various components can be co-jointlyemployed in the methods and articles (e.g., compositions and apparatusesincluding device and methods). For example, the term “comprising” willbe understood to imply the inclusion of any stated elements or steps butnot the exclusion of any other elements or steps.

In general, any of the apparatuses and methods described herein shouldbe understood to be inclusive, but all or a sub-set of the componentsand/or steps may alternatively be exclusive, and may be expressed as“consisting of” or alternatively “consisting essentially of” the variouscomponents, steps, sub-components or sub-steps.

As used herein in the specification and claims, including as used in theexamples and unless otherwise expressly specified, all numbers may beread as if prefaced by the word “about” or “approximately,” even if theterm does not expressly appear. The phrase “about” or “approximately”may be used when describing magnitude and/or position to indicate thatthe value and/or position described is within a reasonable expectedrange of values and/or positions. For example, a numeric value may havea value that is +/−0.1% of the stated value (or range of values), +/−1%of the stated value (or range of values), +/−2% of the stated value (orrange of values), +/−5% of the stated value (or range of values), +/−10%of the stated value (or range of values), etc. Any numerical valuesgiven herein should also be understood to include about or approximatelythat value, unless the context indicates otherwise. For example, if thevalue “10” is disclosed, then “about 10” is also disclosed. Anynumerical range recited herein is intended to include all sub-rangessubsumed therein. It is also understood that when a value is disclosedthat “less than or equal to” the value, “greater than or equal to thevalue” and possible ranges between values are also disclosed, asappropriately understood by the skilled artisan. For example, if thevalue “X” is disclosed the “less than or equal to X” as well as “greaterthan or equal to X” (e.g., where X is a numerical value) is alsodisclosed. It is also understood that the throughout the application,data is provided in a number of different formats, and that this data,represents endpoints and starting points, and ranges for any combinationof the data points. For example, if a particular data point “10” and aparticular data point “15” are disclosed, it is understood that greaterthan, greater than or equal to, less than, less than or equal to, andequal to 10 and 15 are considered disclosed as well as between 10 and15. It is also understood that each unit between two particular unitsare also disclosed. For example, if 10 and 15 are disclosed, then 11,12, 13, and 14 are also disclosed.

Although various illustrative embodiments are described above, any of anumber of changes may be made to various embodiments without departingfrom the scope of the invention as described by the claims. For example,the order in which various described method steps are performed mayoften be changed in alternative embodiments, and in other alternativeembodiments one or more method steps may be skipped altogether. Optionalfeatures of various device and system embodiments may be included insome embodiments and not in others. Therefore, the foregoing descriptionis provided primarily for exemplary purposes and should not beinterpreted to limit the scope of the invention as it is set forth inthe claims.

The examples and illustrations included herein show, by way ofillustration and not of limitation, specific embodiments in which thesubject matter may be practiced. As mentioned, other embodiments may beutilized and derived there from, such that structural and logicalsubstitutions and changes may be made without departing from the scopeof this disclosure. Such embodiments of the inventive subject matter maybe referred to herein individually or collectively by the term“invention” merely for convenience and without intending to voluntarilylimit the scope of this application to any single invention or inventiveconcept, if more than one is, in fact, disclosed. Thus, althoughspecific embodiments have been illustrated and described herein, anyarrangement calculated to achieve the same purpose may be substitutedfor the specific embodiments shown. This disclosure is intended to coverany and all adaptations or variations of various embodiments.Combinations of the above embodiments, and other embodiments notspecifically described herein, will be apparent to those of skill in theart upon reviewing the above description.

What is claimed is:
 1. A removable orthodontic device comprising: aplurality of tooth receiving cavities configured to receive a pluralityof teeth and to exert one or more orthodontic repositioning forces onthe plurality of teeth; and at least one biosensor system comprising: abioreceptor configured to cause a first interaction with one or moretooth motion biomarkers in fluid in the oral cavity, the firstinteraction being related to a first biomarker expression change of theone or more tooth motion biomarkers, and the first biomarker expressionchange being associated with a specific phase of tooth movement of theplurality of teeth; and a biotransducer coupled to the tooth motionbioreceptor, the biotransducer configured to transduce the firstinteraction into a first interaction signal representative of the firstinteraction.
 2. The removable orthodontic device of claim 1, wherein thespecific phase of tooth movement corresponds to velocity of rootmovement of roots of the plurality of teeth.
 3. The removableorthodontic device of claim 2, wherein the velocity of root movement issubstantially zero, thereby indicating a conclusion of a portion of anorthodontic treatment plan.
 4. The removable orthodontic device of claim1, wherein: the bioreceptor is configured to cause a second interactionwith one or more compliance biomarkers, the second interaction beingrelated to a second biomarker expression change associated withcompliance by the patient; and the biotransducer is configured totransduce the second interaction into a second interaction signalrepresentative of the second interaction.
 5. The removable orthodonticdevice of claim 1, wherein the at least one biosensor system includes aprocessor configured to process the first interaction signal into sensordata used to provide one or more recommendations to change the removableorthodontic appliance at a specified time.
 6. The removable orthodonticdevice of claim 1, wherein the at least one biosensor system comprisesone or more of memory, a power source, a communication unit, and anantenna.
 7. The removable orthodontic device of claim 1, furthercomprising a biosensor housing formed from at least a portion of theplurality of tooth receiving cavities and being configured to physicallycouple the bioreceptor to at least one tooth of the plurality of tooth.8. The removable orthodontic device of claim 1, wherein the bioreceptorcomprises a protein that selectively binds to the tooth motionbiomarker.
 9. The removable orthodontic device of claim 1, wherein thebioreceptor comprises one or more of a primary and/or a secondaryprotein.
 10. The removable orthodontic device of claim 1, wherein thebioreceptor comprises an enzyme that selectively acts on the toothmotion biomarker.
 11. The removable orthodontic device of claim 1,wherein the fluid comprises saliva.
 12. The removable orthodontic deviceof claim 11, wherein the tooth movement biomarker is one of:Calgranulin-B, Serum albumin precursor, Immunoglobulin J chain, Igalpha-1 chain C region, Cysteine-rich secretory protein 3 precursor(CRISP-3), Hemoglobin subunit beta, Stratifin, and soluble RANK Ligand(sRANKL).
 13. The removable orthodontic device of claim 1, wherein thefluid comprises gingival crevicular fluid (GCF).
 14. The removableorthodontic device of claim 13, wherein the tooth movement biomarker isone of: prostaglandin E2, Substance P, epidermal growth factor,transforming growth factor, Receptor activator of nuclear factor kappa-Bligand (RANKL), Granulocyet macrophage colony stimulation factor, α2microglobulin, Interleukin 1β, Myeloperoxidase, hyaluronic acid andChondroitin sulfate.
 15. The removable orthodontic device of claim 1,wherein the at least one biosensor system comprises a referenceelectrode comprising the biotransducer material but not a bioreceptormaterial.
 16. The removable orthodontic device of claim 1, wherein thebiosensor comprises one or more of nanowires and nanoparticles.
 17. Theremovable orthodontic device of claim 1, wherein the biosensor comprisesa conductive ink and an elastomeric binder.
 18. The removableorthodontic device of claim 17, wherein the elastomeric bindercomprises: silicone, fluorine rubber, polyurethane and isoprene blockco-polymers.
 19. The removable orthodontic device of claim 1, whereinthe biosensor further comprises a conductive substrate to which thebiotransducer is attached and an insulating layer.
 20. The removableorthodontic device of claim 1, wherein the at least one biosensor systemis one of a plurality of biosensor systems operably coupled to differentportions of the removable orthodontic device.
 21. The removableorthodontic device of claim 1, further comprising a physical sensoroperatively coupled to the removable orthodontic device, wherein thephysical sensor is configured to sense one or more of a force, apressure, a temperature, or a movement of the plurality of teeth. 22.The removable orthodontic device of claim 1, further comprising acommunication module configured to transmit the sensor data to a remotedevice.
 23. A method comprising: receiving a plurality of teeth in anoral cavity of a patient with a plurality of tooth receiving cavities ofa removable orthodontic aligner, the plurality of tooth receivingcavities configured to exert one or more orthodontic repositioningforces on the plurality of teeth; placing a bioreceptor in contact withfluid in the oral cavity to cause a first interaction with one or moretooth motion biomarkers in the fluid, the first interaction beingrelated to a first biomarker expression change of the one or more toothmotion biomarkers, and the first biomarker expression change beingassociated with a specific phase of tooth movement of the one or moreteeth; transducing the first interaction into a first interaction signalrepresentative of the first interaction; and providing the firstinteraction signal.
 24. A removable orthodontic device comprising: meansfor receiving a plurality of teeth in an oral cavity of a patient and toexert one or more orthodontic repositioning forces on the plurality ofteeth; and at least one biosensor system comprising: means for causing afirst interaction with one or more tooth motion biomarkers in fluid inthe oral cavity, the first interaction being related to a firstbiomarker expression change of the one or more tooth motion biomarkers,and the first biomarker expression change being associated with aspecific phase of tooth movement of the plurality of teeth; and meansfor transducing the first interaction into a first interaction signalrepresentative of the first interaction.